Antifungal Activity of Lavandula angustifolia Essential Oil against Candida albicans: Time-Kill Study on Pediatric Sputum Isolates.

The aim of our study was to determine the susceptibility of 15 Candida albicans sputum isolates on fluconazole and caspofungin, as well as the antifungal potential of Lavandula angustifolia essential oil (LAEO). The commercial LAEO was analyzed using gas chromatography-mass spectrometry. The antifungal activity was evaluated using EUCAST protocol. A killing assay was performed to evaluate kinetics of 2% LAEO within 30 min treatment. LAEO with major constituents' linalool (33.4%) and linalyl acetate (30.5%) effective inhibited grows of C. albicans in concentration range 0.5-2%. Fluconazole activity was noted in 67% of the isolates with MICs in range 0.06-1 µg/mL. Surprisingly, 40% of isolates were non-wild-type (non-WT), while MICs for WT ranged between 0.125-0.25 µg/mL. There were no significant differences in the LAEO MICs among fluconazole-resistant and fluconazole-susceptible sputum strains (p = 0.31) and neither among caspofungin non-WT and WT isolates (p = 0.79). The 2% LAEO rapidly achieved 50% growth reduction in all tested strains between 0.2 and 3.5 min. Within 30 min, the same LAEO concentration exhibited a 99.9% reduction in 27% isolates. This study demonstrated that 2% solution of LAEO showed a significant antifungal activity which is equally effective against fluconazole and caspofungin susceptible and less-susceptible strains.

Epidemiology of onychomycosis in Serbia: a laboratory-based survey and risk factor identification.

Onychomycosis is a fungal infection of the fingernails and/or toenails caused by dermatophytes, yeasts and non-dermatophyte moulds. The epidemiology of onychomycosis in Serbia is yet to be fully established. This epidemiological study was aimed at evaluating the epidemiology of onychomycosis in a sample of the Serbian patients at risk of onychomycosis, to determine the fungal aetiological agents and to identify the possible risk factors. The study population included 374 patients from six centres in Serbia with suspected onychomycosis. Demographic data, data about comorbidities, lifestyle, clinical aspects of onychomycosis, trauma, excessive perspiration and personal and family history of previous onychomycosis were studied. Laboratory confirmation of diagnosis was done by direct microscopy, fungal culture and PCR. Diagnosis of onychomycosis was confirmed in 50.8% of patients, who tested positive to at least one laboratory test (direct microscopy, fungal culture or PCR). Trichophyton rubrum was predominant both on toenails (85.98%) and on fingernails (38.46%). Independent risk factors for onychomycosis were: old age (OR = 2.285; P < 0.001), family history of previous onychomycosis and/or tinea pedis (OR = 2.452; P = 0.005), excessive perspiration (OR = 2.165; P = 0.002) and higher degree of hyperkeratosis (OR = 1.755; P = 0.020). This is a first epidemiological study of onychomycosis from Serbia.

Host-Pathogen Interaction and Resistance Mechanisms in Dermatophytes.

Dermatophytes are widely distributed in the environment, with an estimated prevalence of 20-25% of the the global population yearly. These fungi are keratinophilic and keratinolytic and cause the infection of keratin-rich structures such as skin, hair, and nails. The pattern of this infectious disease covers a wide spectrum from exposed individuals without symptoms to those with acutely inflammatory or non-inflammatory, chronic to invasive, and even life-threatening symptoms. This review summarizes current information on the pathogenicity, virulence factors, and drug resistance mechanisms associated with dermatophytes. A greater number of virulence factors of these fungi are important for the occurrence of infection and the changes that occur, including those regarding adhesins, the sulfite efflux pump, and proteolytic enzymes. Other virulence factors include mechanisms of evading the host defense, while the development of resistance to antifungal drugs is increasing, resulting in treatment failure. The investigation of host-pathogen interactions is essential for developing a more complete understanding of the mechanisms underlying dermatophyte pathogenesis and host response to inform the use of diagnostics methods and antifungal therapeutics to minimize the high fungal burden caused by dermatophytes and to control the spread of resistance.

Human infections by Hymenolepis diminuta in Europe: a case report and literature review.

We performed a review of published and gray literature of human Hymenolepis diminuta cases across Europe up to July 2022. Of all detectable publications and records, we further analyzed only those that contained demographic, clinical or epidemiological data regarding the infected subjects. Additionally, one case of hymenolepiasis in a 16-mo-old boy living in the urban part of Belgrade was included in the analysis. Published studies were based in 13/50 European countries and identified 104 laboratory-confirmed cases in total. Almost one-half (49%) of all cases originated from Mediterranean countries. Among symptomatic children, the infection most often manifested with diarrhea, abdominal pain, allergic reaction and behavioral changes. The diagnosis was made by the detection and identification of H. diminuta eggs or parts of strobila in stool samples, although cases of misdiagnosis were also reported. The parasite clearance was established with praziquantel or niclosamide, while the administration of albendazole or mebendazole led to unfavorable results. Future multicentric prospective studies focused on infection screening and the gathering of detailed sociodemographic and clinical data could provide an updated insight into the true distribution and characteristics of H. diminuta infection across Europe.

Fungal contamination of medical students' mobile phones from the University of Belgrade, Serbia: a cross-sectional study.

The study aimed to characterize fungal contamination of medical students' mobile phones, investigate mobile phones' usage and cleaning habits, identify independent risk factors for fungal contamination, and awareness of mobile phones as a potential route of infection. In a cross-sectional study, medical students' mobile phones were sampled for possible fungal contamination. The questionnaire was used to record mobile phone usage, cleaning habits, and awareness of mobile phones as a source of infection. A total of 492 medical students were included and fungal contamination of mobile phones was confirmed in 32.11%. The most frequent fungal isolates on students' mobile phones were Candida albicans (28.5%), followed by Aspergillus niger (11.4%), and Penicillium chrysogenum (9.5%). Factors independently associated with fungal contamination of students' mobile phones were: lack of mobile phone cleaning (OR = 0.381; p < 0.001), and usage of mobile phones near patients' beds (OR = 0.571; p = 0.007). The results of this study confirmed that students who use their mobile phones in hospital wards have a higher rate of fungal contamination. The development of active surveillance and preventive strategies is needed to reduce the risk of cross-contamination and increase awareness of fungal transmission via mobile phones.

Correlation of clinical characteristics, by calculation of SCIO index, with the laboratory diagnosis of onychomycosis.

INTRODUCTION: Onychomycosis is a chronic fungal infection with increasing incidence and the global prevalence is estimated to be 5.5%. The aim of our study was to perceive objectively severity of onychomycosis by calculating Scoring Clinical Index for Onychomycosis and to correlate this index with accurate laboratory diagnosis in our patients. MATERIALS AND METHODS: The study population comprised of 417 patients with laboratory confirmed onychomycosis. For each patient, we recorded basic demographic information, site of infection, the most affected nail with onychomycosis, clinical presentation, and type of onychomycosis. The evaluation of the disease severity was based on Scoring Clinical Index for Onychomycosis which was calculated for every patient separately. Mycological identification was done by microscopy and fungal culture. RESULTS: The majority of patients had distal and lateral subungual onychomycosis (95.44%) that was localized on big toe (62.59%), with female to male ratio 1.24:1. Male patients had significantly more nails affected with onychomycosis compared with female patients (p = 0.011), while female had significantly more often onychomycosis on fingernails 2-5 (p < 0.05), and they reported significantly more often pain (p < 0.05) and esthetic problems (p < 0.05). Mean Scoring Clinical Index for Onychomycosis was 16.76. Dermatophytes were most frequently isolated (91.85%). In patients with onychomycosis caused by dermatophytes, Scoring Clinical Index for Onychomycosis had significantly higher values (p = 0.032). CONCLUSION: Comprehensive understanding of disease characteristics will allow introduction of individualized treatment plan for each patient, based on proper fungal identification and standardized method of evaluating disease severity, which could help the patient achieve a complete cure.

Diagnosis of Trichomonas vaginalis infection: The sensitivities and specificities of microscopy, culture and PCR assay.

OBJECTIVES: The aim of this study was to compare wet mount-, Giemsa stain-, acridine orange fluorescent stain-, cultivation- and polymerase chain reaction (PCR)-based approaches to establish which method or combination of methods was most effective in the laboratory diagnosis of trichomoniasis. STUDY DESIGN: Out of 200 investigated patients with various gynecological complaints, Trichomonas vaginalis infection was detected in 27 (13.5%) by any of methods investigated. Among women with trichomonads, a typical clinical finding was presented in only nine. For analysis of sensitivity and specificity of the methods used, the receiver operating characteristic (ROC) curve concept with culture as a gold standard was applied. RESULTS: Infection was diagnosed by wet mount in 14 (7.0%) women, by Giemsa stain in 11 (5.5%) and by acridine orange stain in 16 (8.0%) women. In 21 (10.5%) women, it was diagnosed by culture in Diamond's medium, and in 22 (11.0%) by PCR. For the initial diagnosis of trichomoniasis, wet preparation is the test that is widely available in most STD clinics, but its sensitivity is poor (66.67%). Giemsa stain shows a low sensitivity of 52.38%. Acridine orange shows reasonable sensitivity and specificity of 71.43% and 99.44%, respectively. The sensitivity and specificity of PCR (80.95% and 97.21%) did not exceed that of culture. CONCLUSION: With regard to the fact that trichomoniasis can have an atypical or even asymptomatic course, in order to accurately diagnose this disease, microbiological investigation is necessary. Comparison of different methods showed that at least two techniques, such as culture and acridine orange staining, have the potential for better diagnosis of T. vaginalis infection. PCR detection of infection has been demonstrated to be highly specific and sensitive, but its availability and cost effectiveness are in question. PCR could provide an alternative for laboratory diagnosis of trichomoniasis by culture.

Fatal Strongyloides stercoralis infection in a young woman with lupus glomerulonephritis.

We report a case of fatal Strongyloides stercoralis (S. stercoralis) infection in a 35-year-old female with lupus glomerulonephritis after prolonged steroid therapy. An epidemiological evaluation revealed that our patient originated from a rural area in Bosnia and Herzegovina, which was an endemic region of S. stercoralis infection in the former Yugoslavia. She had severe gastrointestinal and pulmonary symptoms and history of a 13 kg weight loss in 3 months. Histopathological examination of large bowel mucosa showed nematode larvae. Microscopy of stool, sputum and urine samples confirmed S. stercoralis. The diagnosis was delayed because of the low suspicion index, the absence of eosinophilia and non-specific signs of infection, which could be a result of the underlying disease or the effects of corticosteroids. This case highlights the importance of screening for S. stercoralis in patients starting immunosuppressive therapy, especially if they are from endemic areas. Timely diagnosis and therapy could save the patient's life.

Human case of fasciolosis in Serbia treated with triclabendazole.

INTRODUCTION: The number of humans infected by Fasciola hepatica is increasing worldwide. Humans can become accidental hosts by ingesting drinking water or plants contaminated with metacercariae. CASE REPORT: We reported a case of a 68-year-old Serbian woman, in which the diagnosis of acute fasciolosis had been established after serious diagnostic concerns. Based on clinical picture (episodic right upper quadrant abdominal pain, febrility and generalized body pain) and biochemical analyses (high eosinophilia and high activity of alkaline phosphatase), she was appointed as suspected to the acute fasciolosis. Stool and duodenal aspirate exams were negative for Fasciola ova. In the absence of adequate serologic diagnostic for fasciolosis in Serbia, the diagnosis was confirmed using enzyme immunoassays and immunoblot at the Institute for Tropical Diseases in Hamburg, Germany. Soon after triclabendazole was administered, the symptoms disappeared and biochemical values returned to normal. CONCLUSION: The diagnosis of human fasciolosis may be problematic and delayed, especially in non endemic areas, because physicians rarely encounter this disease and a long list of other diseases must be considered in the differential diagnosis. The syndrome of eosinophilia, fever, and right upper quadrant abdominal pain suggest acute fasciolosis. Unclear source does not rule out fasciolosis.

Human external ophthalmomyiasis caused by Lucilia sericata Meigen (Diptera: Calliphoridae)--a green bottle fly.

Ophthalmomyiasis externa is the result of infestation of the conjunctiva by the larval form or maggots of flies from the order Diptera. If not recognized and managed appropriately, it can be complicated by the potentially fatal condition ophthalmomyiasis interna. Ophthalmomyiasis externa is mainly caused by the sheep bot fly (Oestrus ovis). We present the first case, to our knowledge, of ophthalmomyiasis externa in an elderly woman from Belgrade caused by Lucilia sericata Meigen--a green bottle fly.

[Detection of laboratory biomarkes in haematological and pulmonology patients at high risk for aspergillosis].

INTRODUCTION: During the past decades a dramatic increase in the incidence of invasive fungal diseases, especially invasive aspergillosis has been observed. Undiagnosed and diagnosed late invasive aspergillosis is followed by lethality of up to 90%. Detection of early laboratory biomarkers (galactomannan and anti-Aspergillus antibodies) contributes to early diagnosis and is used for screening, as well as for monitoring therapy of aspergillosis. OBJECTIVE: The aim was to evaluate usefulness of "non-culture" methods (galactomannan and anti-Aspergillus antibodies IgA, IgM and IgG) for early diagnosis of aspergillosis in high-risk patients. METHODS: Prospective two-year study involved 262 high-risk patients for aspergillosis. In pulmonology and haematology patients (adults and children) blood samples were tested on galactomannan and anti-Aspergillus antibodies. RESULTS: Early laboratory biomarkers were statistically significantly higher in pulmonology patients (p = 0.00033). However, in haematological patients galactomannan was a more frequently positive finding, while in pulmonology patients it was the finding of anti-Aspergillus antibodies. It is interesting that, despite the possible immunosuppression, in about 1/3 of haematological patients anti-Aspergillus antibodies were confirmed. CONCLUSION: Early diagnosis and treatment of aspergillosis represent both clinical and laboratory problem. Nowadays, the application of new "non-culture" methods is of particular importance for the outcome of aspergillosis. Clinical features, laboratory findings of biomarkers and their correct interpretation significantly increase the possibility of timely implementation of appropriate therapy. In this regard, the new organization of reference laboratory for medical mycology has significantly improved the outcome of aspergillosis in high-risk patients in our country. However, further investigations, implementation of European standards and introduction of new diagnostic methods are necessary in this field.

[Antimycotics susceptibility testing of dermatophytes].

Dermatophytes are moulds that produce infections of the skin, hair and nails of humans and animals. The most common forms among these infections are onychomycosis and tinea pedis affecting 20% of world population. These infections are usually chronic. The treatment of dermatophytoses tends to be prolonged partly because available treatments are not very effective. Antifungal drug consumption and public health expenditure are high worldwide, as well as in Serbia. For adequate therapy, it is necessary to prove infection by isolation of dermatophytes and to test the antifungal susceptibility of isolates. Susceptibility testing is important for the resistance monitoring, epidemiological research and to compare in vitro activities of new antifungal agents. The diffusion and dilution methods of susceptibility tests are used, and technical issues of importance for the proper performance and interpretation of test results are published in the document E.DEF 9.1 (EUCAST) and M38-A2 (CLSI). The aim of our paper is to promptly inform the public about technical achievements in this area, as well as the new organization of laboratory for medical mycology in our country. The formation of laboratory networks coordinated by the National Reference Laboratory for the cause of mycosis need to enable interlaboratory studies and further standardization of methods for antifungal susceptibility testing of dermatophytes, reproducibility of tests and clinical correlation monitoring (MIK values and clinical outcome of dermatophytosis). The importance of the new organization is expected efficient improvement in the dermatophytosis therapy at home, better quality of patient's life and the reduction of the cost of treatment.

[IgG serum antibody responses in suspected liver cystic echinococcosis patients].

INTRODUCTION: Cystic echinococcosis is a parasitic zoonosis caused by a tapeworm Echinococcus granulosus. Liver infection is the most common form of human echinococcosis. Diagnosis of liver echinococcosis could be made by different methods. It is very important to have good anamnesis and epidemiological evaluation, clinical status and clinical investigations such as X-ray examination, ultrasonography, nuclear magnetic resonance, computerized tomography, laboratory analysis and serology. Different serologic tests could be used for diagnosis of echinococcosis: complement fixation test, immunoelectrophoresis, indirect haemagglutination, latex agglutination, indirect fluorescence test and enzyme immunoassays sush as ELISA test. OBJECTIVE: The aim of this study was to investigate anti-E granulosus antibodies by ELISA test as a complementary laboratory method in the diagnosis of human echinococcosis. METHOD: In this study, we investigated 212 patients of suspected liver cystic echinococcosis. ELISA test was used for detection of whole anti-E granulosus IgG antibodies specific for AgB. RESULTS: Out of 212 patients, in 26 (12.26%) patients echinococcosis was confirmed by ELISA test. In 7/26 patients who had data about the ultrasonographic morphological type of cyst according to Gharbi, high values of seropositivity were found. Borderline values were found in 15/212 (7.08%) patients. In three patients with borderline values, second sera samples were examined after 18-22 days. In one patient, the test showed the same value, in the second patient echinococcosis was confirmed while in the third, the test showed a negative value. In 171 (80.66%) patients, anti-Echinococcus antibodies were not found by ELISA test. CONCLUSION: It has already been known that serological investigation has some limitations, but detection of specific antibodies remains an indispensable mark in evaluation of suspected liver cystic echinococcosis patients.

[Antifungal drug resistance: mechanisms of resistance, freqeuncy, prevention and control of resistance].

Fungi are important causes of human infections, especially systemic mycoses. Pathogenic fungi have many complex mechanisms of resistance to antifungal drugs. In this article, information about the cellular, genetic and clinical factors contributing to antifungal-drug resistance, and their diagnostic and epidemiologic characteristics are reviewed. Understanding the mechanisms of resistance should assist in developing better detection and preventive strategies. The emergence of acquired antifungal resistance, especially in immunocompromised hosts, has necessitated routine antifungal susceptibility testing. Initial antifungal screening of clinical isolates by the disk-diffusion method would be followed by confirmation of resistant strains by the broth dilution method. Also, strategies to avoid and suppress the antifungal resistance are discussed. There are several strategies to overcome antifungal resistance, including increased antifungal-dose intensity, immunomodulation, combined antifungal therapy and new antifungals.

[Characteristics of the immune response in protozoan infections]. / Karakteristike imunskog odgovora u toku infekcije protozoama.

INTRODUCTION: When protozoa enter the blood stream or tissues they can often survive and replicate because they adapt to the resisting natural host defenses. The interaction of immune system with infectious organisms is a dynamic interplay of host mechanisms aimed at eliminating infections and microbial strategies designed to permit survival in the face of powerful effector mechanisms. Protozoa cause chronic and persistent infections, because natural immunity against them is weak and because protozoa have evolved multiple mechanisms for evading and resisting specific immunity. NATURAL AND SPECIFIC IMMUNE RESPONSE TO PROTOZOA: Different protozoa vary greatly in their structural and biochemical properties and stimulate distinct patterns of immune responses and have evolved unique mechanisms for evading specific immunity. Protozoa activate quite distinct specific immune responses, which are different from the responses to fungi, bacteria and viruses. Protozoa may be phagocytozed by macrophages, but many are resistant to phagocytic killing and may even replicate within macrophages. T. brucei gambiense is the best example of protozoa which can induce humoral immune response because of its extra-cellular location. In Leishmania sp. infections, cellular defense mechanisms depend upon CD4+ T-lymphocytes and activate macrophages as effector cells that are regulated by cytokines of Th1 subset. Plasmodium sp. is a protozoa which show the diversity of defence mechanisms which can be cellular or humoral, depending on Ag and protozoa's location. IMMUNE EVASION MECHANISMS OF PROTOZOA: Different protozoa have developed remarkably effective ways of resisting specific immunity: a) anatomic sequestration is commonly observed with protozoa Plasmodium and T. gondii; b) some protozoa can become resistant to immune effector mechanisms: Trypanosoma, Leishmania and T. gondii; c) some protozoa have developed effective mechanisms for varying their surface antigens: Plasmodium and Trypanosoma; d) some protozoa shed their antigen coats, either spontaneously or after binding with specific antibodies: E. histolytica; e) some protozoa alter host immune response by nonspecific and generalized immunosuppression (abnormalities in cytokine production, deficient T cell activation): Trypanosoma, Leishmania, Toxoplasma, Entamoeba. CONCLUSION: Protozoa activate numerous, different immune mechanisms in human body. Evolution, progression and outcome of diseases depend upon these mechanisms. Resent progresses in research have defined and selected Ag as candidates for new vaccines. Better definitions regarding the role of cytokines in protozoan infections will facilitate rational development of cytokines and cytokine antagonists and their use as immunotherapeutic agents.

Correlation between clinical symptoms and diagnosis of trichomoniasis in women.

The aim of this study was to investigate correlation between clinical symptoms and diagnosis of trichomoniasis in women. 200 women were included in the study. Swabs were taken from all patients from the posterior vaginal fornix. Each sample was examined using the following five methods: wet mount, Giemsa stain, acridine orange fluorescence stain, cultivation in Diamond medium and PCR method. Trichomoniasis was diagnosed in 27 women using any of the applied methods and 33.3% presented with typical frothy yellow-green discharge, characteristic for tichomoniasis and yellowish discharge characteristic for the third group of vaginal discharge. White discharge, characteristic for Candida infection, was found in 18.5% of patients with diagnosed trichomoniasis. Finally, 14.8% of positive patients had a normal discharge. Based on the results of our study we came to the conclusion that microbiological investigations are necessary for accurate diagnosis of trichomoniasis, as well as for revealing asymptomatic infections, in order to prevent spreading of this relatively common disease.

[Human babesiosis--recent discoveries]. / Humana babezioza--nova saznanja.

INTRODUCTION: Babesiosis is caused by intraerythrocytic parasites of the genus Babesia, which is a common animal infection worldwide. This protozoa requires both a competent vertebrate and a nonvertebrate host (Ixodes sp. etc.) to maintain the transmission cycle. HUMAN BABESIOSIS: Human babesiosis is predominantly caused by Babesia microti (rodent-borne piroplasm, an emerging zoonosis in humans in North America) and by Babesia divergens (bovine pathogen, in Europe). Occasionally, infection in America is caused also by a newly recognized species, so-called WA1 piroplasm. The spectrum of human babesiosis in the USA is broad, and ranges from an apparently silent infection to a fulminant. In Europe, babesiosis is considerably rarer, but more lethal (42% mortality rate in Europe and 5% in the USA, for clinically apparent infections) and mostly in splenectomized patients. Various determinants are involved in the severity of infection, such as age, immunocompetence and coinfection with other pathogens (Borrelia burgdorferi). B. microti antigens can trigger specific activation of T-cells and the infection can be effectively controlled by a Th1-dominant CD4+ T-cell response. The diagnosis of babesiosis should include examination of blood smears stained by Giemsa, as well as serologic evaluation with indirect immunofluorescent antibody tests and possibly PCR. The treatment of babesiosis depends on severity of cases; if it is mild it resolves spontaneously, whereas very severe cases with B. divergens require prompt treatment that includes erythrocyte exchange transfuision along with intravenous clindamycin and oral quinine to arrest hemolysis and prevent renalfailure. This paper offers an overview of recent developments in the investigation of Babesia sp. and babesiosis.

[First case of Malassezia globosa isolation in Serbia]. / Prvi slucaj isolacije Malassezia globosa kod nas.

Today is known that genus Malassezia includes seven species: M. furfur, M. sympodialis, M. obtusa, M. globosa, M. restricta, M. sloofflae and M. pachydermatis, but role of each of the species in the pathogenesis of disease has not been elucidated yet, so further laboratory isolation and identification are necessary. We report the first case of isolation of Malassezia globosa in Serbia (Belgrade), in a patient suffering from Pityriasis versicolor. Identification of M. globosa was based on macroscopic, microscopic and biochemical characteristics. Isolation was done on Leeming and Notman medium and on mDixona agar, at 350C, during 7 days in aerobic conditions. Also the yeast's biochemical phenotype was determined as catalase (+), lipase (+), esculin degradation (-), Tween (20, 40, 60 and 80) assimilation (-). M. globosa is a lipophilic yeast of the genus Malassezia and the common member of the skin flora. In concordance with some predisponing factors M. globosa is implicated in the pathogenesis of several skin diseases (pityriasis versicolor, malassezia foliculitis, seborheic dermatitis and some forms of atopic dermatitis). In immunocompromised patients and neonates this yeast can even cause fatal systemic infections. Because the role of Malassezia spp. In pathogenesis of skin disease is not still determined, we suggest laboratory diagnosis and identification of these species as a routine diagnostic procedure.