Essential Oils of Three Balkan Abies Species: Chemical Profiles, Antimicrobial Activity and Toxicity toward Artemia salina and Drosophila melanogaster.

This study focused on the essential oils (EOs) isolated from needles with twigs of three indigenous Balkan Abies species (A. alba, A. × borisii-regis and A. cephalonica) regarding their chemical composition, antimicrobial activity and toxicity toward crustaceans and insects. Even though distinct phytochemical profiles of dominant volatiles were revealed for each species, ß-pinene and α-pinene represented the first two major volatiles in all three EOs. Antimicrobial activity of EOs has shown inhibitory effect against all 17 studied strains (ATCC and respiratory isolates) in the range of 0.62-20.00 mg/mL (MICs). Further, all three EOs exhibited strong toxicity (LC50 <100 µg/mL) in Artemia salina lethality bioassay, but with significant differences that depended on the EO type. Additionally, tested EOs have shown a certain level of toxicity against Drosophila melanogaster, mostly at the highest tested concentration (3 %) which caused significant prolongation of developmental time, larvicidal effect and pupal mortality. In the three biological assays performed, there was no observed inhibitory effect or weakest activity for A. alba EO. Further, A. cephalonica EO has shown the highest levels of antimicrobial activity and toxicity toward A. salina, while in relation to the insecticidal potential, A. cephalonica and A. × borisii-regis EOs exhibited similar level of toxicity against D. melanogaster.

Essential Oils of Six Achillea Species: Chemical Profiles, Antimicrobial Potential and Toxicity toward Crustaceans.

This is the first comparative study on antimicrobial activity of essential oils (EOs) of six Achillea species (A. crithmifolia, A. distans, A. grandifolia, A. millefolium, A. nobilis, and A. lingulata) against ATCC and clinical microbial strains isolated from human swabs and on their toxicological potential on crustaceans. Oxygenated monoterpenes represented the major compound class in all six EOs, ranging from around 50 % (A. millefolium, A. nobilis, and A. lingulata) up to over 80 % (A. crithmifolia and A. grandifolia). Specific phytochemical profiles of the major volatiles were determined for each species. Still, borneol was the dominant compound in EOs of three yarrow species: A. distans, A. lingulata, and A. millefolium. Other major compounds were 1,8-cineole in A. crithmifolia, camphor in A. grandifolia, and artemisia ketone in A. nobilis EO. Antimicrobial activities of EOs were examined by a microdilution method revealing inhibitory action against all 15 studied strains in the range of 0.07-20.00 mg/mL (MICs). High toxicity (LC50 <100 µg/mL) in Artemia salina acute toxicity bioassay was a shared characteristic, appearing in five out of six tested EOs, i. e., those isolated from A. lingulata, A. millefolium, A. distans, A. nobilis, and A. grandifolia. In both types of biological tests, A. lingulata EO has shown the highest and A. crithmifolia EO the lowest potential. The strongest activities, recorded for EO of A. lingulata, which was the only studied species from the section Anthemoideae, may be explained by its somewhat unusual chemical composition. According to the obtained results, the second dominant compound of this EO is γ-palmitolactone, which may be a significant agent regarding the antimicrobial activity and toxicity to crustaceans, either on its own or in union with other components, and it may be a suitable topic for further research.

Pseudotsuga menziesii (Pinaceae): Volatile Profiles, Antimicrobial Activity and Toxicological Evaluation of Its Essential Oil.

The present article investigates the chemical composition of volatiles of essential oil (EO) and headspace (HS) fraction, as well as biological activities of EO obtained from needles with twigs of Pseudotsuga menziesii var. menziesii cultivated in Serbia. The major class of compounds was monoterpene hydrocarbons with α-terpinolene, sabinene and ß-pinene (EO), and sabinene, α-terpinolene and ß-pinene (HS) as the dominant volatiles. Tested EO exhibited mostly low antimicrobial potential against investigated strains (ATCC and respiratory isolates), where MICs ranged 1.25-20.00 mg/mL. Nevertheless, based on presented results, where antimicrobial testing was done for the first time on human respiratory system isolates, there is a potential of this EO to be used as an adjuvant in the treatment of human respiratory infections, especially those caused by Pseudomonas aeruginosa or Candida albicans strains. Regarding toxicological evaluation, EO showed moderate toxicity in Artemia salina toxicity bioassay (LC50 =347.41, after 24 h) as well as week toxicity against Drosophila melanogaster with the ability only to moderately delay larval and pupal development.

Tigecycline susceptibility of multidrug-resistant Acinetobacter baumannii from intensive care units in the western Balkans.

Tigecycline can be effective to treat infections of carbapenem-resistant Acinetobacter baumannii (CRAB) however, no interpretive criteria have been approved so far. The objectives of this study were to evaluate the proportion of CRAB isolates and to compare gradient test with a broth microdilution (BMD) method for tigecycline susceptibility testing of A. baumannii.This study included 349 multidrug-resistant (MDR) Acinetobacter spp. collected from Serbia, Montenegro, Bosnia and Herzegovina in 2016 and 2017. Antibiotic susceptibility testing was performed by disk diffusion, VITEK2, gradient, ComASP Colistin. Tigecycline susceptibilities were interpreted according to breakpoints of European Committee on Antimicrobial Susceptibility Testing (EUCAST) and Food and Drug Administration (FDA).Majority of the tested isolates were CRAB (92.8%). Tigecycline MIC50/MIC90 values were 4/8 µg/mL by BMD and 0.5/4 µg/mL by gradient test. Essential agreement for BMD and gradient test amounted to 65.1%. With EUCAST breakpoints, categorical agreement (CA) was achieved in 38% isolates. Major discordance (MD-false susceptibility/resistance) and minor discordance (mD-false categorization involving intermediate results) were observed in 10% and 57% A. baumannii, respectively. With FDA breakpoints, CA, MD and mD were observed in 44%, 16% and 47% isolates, respectively. Colistin resistance was 2.1%.The study highlights a high proportion of CRAB and several discordances between BMD and gradient test which may lead to inappropriate therapy.

Synthesis, characterization, DFT study, DNA/BSA-binding affinity, and cytotoxicity of some dinuclear and trinuclear gold(III) complexes.

In this study, we have synthesized a series of dinuclear and trinuclear gold(III) complexes of the general formula [Au2(N-N)Cl6] (1-3) for dinuclear and [Au3(N-N)2Cl8]+ (4-6) for trinuclear compounds, respectively, in which N-N is a bidentate ligand (1,4-diaminobutane; 1,6-diaminohexane or 1,8-diaminooctane). These complexes were characterized by elemental analysis, molar conductivity, and spectroscopic techniques (IR, UV-Vis, 1H NMR, ESI-MS). We performed DFT calculations to get insight into the geometry of the studies complexes. DNA-binding studies were performed by UV-Vis spectrophotometry and fluorescence spectroscopy. The results of competitive reactions between gold(III) complexes and ethidium bromide (EB) towards DNA have shown that selected complexes can displace EB from DNA-EB adduct. In addition, these experiments confirm that polynuclear gold(III) complexes interact with DNA covalently or via intercalation. Furthermore, high values of binding constants of gold(III) complexes towards bovine serum albumin (BSA) protein indicate good binding affinity. In addition, redox stability of complexes in the presence of DNA/BSA was confirmed by cyclic voltammetry. Results of the interactions between gold(III) complexes with DNA/BSA were discussed in reference to molecular docking data obtain by Molegro virtual docker. The cytotoxic activity of synthesized gold(III) complexes was evaluated on human breast cancer cell line (MDA-MB-231), human colorectal cancer cell line (HCT-116), and normal human lung fibroblast cell line (MRC-5). All complexes dose-dependently reduced cancer and normal cells viabilities, with significant cytotoxic effects (IC50 < 25 µM) for trinuclear gold(III) complexes (4, 5) on HCT-116 cells.

A pilot study in Serbia by European Clostridium difficile Infection Surveillance Network.

Clostridium (Clostridioides) difficile infections (CDIs) are among the most frequent healthcare-associated infections in Serbia. In 2013, Serbia participated in the European Clostridium difficile Infection Surveillance Network (ECDIS-Net) who launched a pilot study to enhance laboratory capacity and standardize surveillance for CDI. Two clinics of Clinical Center of Serbia [Clinic for Infectious and Tropical Diseases (CITD) and Clinic of Orthopedic Surgery and Traumatology (COT)] from Belgrade and one general hospital from another metropolitan area of Serbia, Uzice, participated. During a period of 3 months in 2013, all patients with diagnosed CDI were included. The CDI incidence rates in CITD, COT, and General Hospital Uzice were 19.0, 12.2, and 3.9 per 10,000 patient-days, respectively. In total, 49 patients were enrolled in the study with average age of 72 years. A complicated course of CDI was found in 14.3% of all patients. Six (12.2%) of 49 patients died, but not attributable to CDI. Of 39 C. difficile isolates, available for ribotyping, 78.9% belonged to ribotype 027; other PCR ribotypes were 001, 015, 002, 005, 010, 014, and 276. Antimicrobial susceptibility testing revealed low levels of MIC50 and MIC90 for metronidazole (0.5 µg/ml both) and vancomycin (0.25 and 0.5 µg/ml), while 28 strains of ribotype 027 were resistant to moxifloxacin with MIC ≥4 µg/ml. National surveillance is important to obtain more insight in the epidemiology of CDI and to compare the results with other European countries. This study by ECDIS-Net gives bases for a national surveillance of CDI in Serbia.

The Relevance of Osteoclastic and Osteoblastic Activity Markers Follow-Up in Patients on Antiresorptive Osteoporosis Treatment.

In general, markers of bone formation and markers of bone resorption are changing synergistically, so the monitoring of any osteoclastic and any osteoblastic marker should reflect the rate of bone transformation. The aim of the study is to monitor the bone metabolism markers in postmenopausal women with osteoporosis and osteopenia along with the variations caused by the effects of bisphosphonate therapy. The study involved 55 women of average age of 57.95 years, with osteopenia or osteoporosis. The patients with osteoporosis were treated with bisphosphonates (75 mg once a week); the laboratory tests were performed before the treatment and 6 months later. Patients with osteopenia were evaluated at the first assessment and 6 months later. The tests included bone densitometry, dual-energy X-ray absorptiometry, osteocalcin, alkaline phosphatase, collagen 1 N-terminal pro-peptide (P1NP), and beta C telopeptide of type I collagen (CTX). The mean T-score was -2.80 ± 0.63 before therapy and -2.64 ± 0.45 6 months later (p < 0.001). Women with osteoporosis had elevated levels of osteocalcin and P1NP at the first assessment, whereas the alkaline phosphatase level did not change with the treatment. After the introduction of antiresorptive therapy, the levels of osteocalcin and P1NP significantly decreased (p < 0.001). In the group with osteopenia, the biochemical markers activity were increased in both assessments. In patients with osteoporosis, Beta-CTX was increased in the first evaluation, and decreased after treatment (p = 0.001). The results indicate that the assessment of biochemical markers of bone metabolism show excellent results in the assessment of prognosis, monitoring the course and the response to various treatment regimens of osteoporosis and evince strong correlation with standard densitometry and dual-energy X-ray absorptiometry procedures. P1NP and CTX show better diagnostic applicability compared with osteocalcin and alkaline phosphatase. The analysis of the activity of biochemical markers may obtain early information on the therapeutic response, before definitive assessment by bone density measurements.

Epicuticular Waxes Provide Insights into Phytochemical Differentiation of Natural Populations of Pinus mugo Turra sensu stricto.

Phytochemical diversity and differentiation of nine native populations of Pinus mugo sensu stricto from Julian Alps, Southern Carpathians and Balkan Peninsula were analyzed in regard to their epicuticular wax compounds. GC/MS and GC-FID analyses of the hexane extracts of needle samples, collected from 118 individuals of P. mugo, revealed the presence of six diterpenes, three primary alcohols and 10 n-alkanes ranging from C19 to C29 (except n-alkane C21 ). According to simple linear regression, the contents of epicuticular wax compounds showed generally weak correlations with tested bioclimatic, orographic and geographic parameters, leading to assumption about their genetic conditioning. The multivariate statistical analyses suggested the existence of two chemical entities: the Alpine and the South Carpathian, while the Balkan populations appeared heterogeneous as three of them belonged to the Alpine and one to the South Carpathian group. The obtained results are largely consistent to those previously published in reference to morpho-anatomical and molecular characters of P. mugo, supporting the hypothesis of colonization of Balkan Peninsula from two different glacial refugia: from the Alps (across the Dinarides) and from the Southern Carpathians. Extending this study to the entire range of P. mugo complex should provide a clearer picture of its phytochemical diversity and differentiation.

Dispersive micro-solid phase extraction of 16 priority polycyclic aromatic hydrocarbons from water by using thermally treated clinoptilolite, and their quantification by GC-MS.

The authors report on a novel sorbent (thermally treated natural zeolite; clinoptilolite) for use in dispersive micro-solid phase extraction (D-µ-SPE) of polycyclic aromatic hydrocarbons (PAHs) from water samples. The method was applied to the D-µ-SPE of 16 priority PAHs which then were quantified by gas chromatography with mass spectrometric detection (GC-MS). The method was validated in terms of specificity and selectivity, linearity and linear range, accuracy, precision, uncertainty, limits of detection and quantification. Figures of merit include (a) linear analytical ranges between 2.08 and 208 ppb, and (b) detection limits in the range from 0.01 to 0.92 ppb. The method was successfully applied to the determination of PAHs in river waters. Graphical abstract Schematic representation of dispersive micro-solid phase extraction (D-µ-SPE) of trace levels of PAHs in water samples by using thermally treated clinoptilolite as sorbent prior to gas chromatography-mass spectrometry analysis (GC-MS).

Occurrence of Clostridium difficile infections in Serbia and high proportion of PCR ribotype 027 strains in two hospitals in Belgrade.

BACKGROUND: The incidence of Clostridium difficile infections (CDI) in the Clinical Center of Serbia (CCS) and the entire Serbia has been constantly rising in the previous 5 years. We aimed to study C. difficile PCR-ribotypes isolated from patients hospitalized at two healthcare institutions: CCS and Specialized Hospital for Cerebrovascular Diseases "Sveti Sava" (SS), both of them from Belgrade, and to investigate the incidence rates of CDI in hospital settings in Serbia, from 2009 to 2013. METHODS: The Bacteriology laboratory database at Clinic for Infectious and Tropical Diseases of CCS was queried from January 1, 2009 to December 31, 2013 for all patients who underwent immunochromatographic toxin A and/or toxin B stool testing and C. difficile stool culture for suspected infection caused by this bacterium. Toxigenic culture was not performed. Ninety- six C. difficile isolates were then selected and characterized by PCR-ribotyping. These were obtained from 94 patients hospitalized in different clinics of CCS and SS from November 2011 to December 2013. RESULTS: Among 6164 stool samples sent to Bacteriology laboratory for culture of C. difficile and toxin detection during the study period, 1775 (28.8%) were positive, displaying linear trend of growth. From 96 isolates, typed by PCR-ribotyping, majority (85; 88.54%) belonged to PCR-ribotype 027. The remaining 11 isolates belonged to PCR-ribotypes 014/020 (3 isolates), 015, SLO 191 (two isolates each), 017, 018, 070 and 001/072 (one isolate each). CONCLUSION: Our results demonstrated that C. difficile PCR-ribotype 027 is by far predominant in two hospital settings in Belgrade, at least since 2011.

Seasonal variations of Saanen goat milk composition and the impact of climatic conditions.

The aim of this research was to investigate the effect of climatic conditions and their impact on seasonal variations of physico-chemical characteristics of Saanen goat milk produced over a period of 4 years. Lactation period (early, mid and late) and year were considered as factors that influence physico-chemical composition of milk. Pearson's coefficient of correlation was calculated between the physico-chemical characteristics of milk (fat, proteins, lactose, non-fat dry matter, density, freezing point, pH, titrable acidity) and climatic condition parameters (air temperature, temperature humidity index-THI, solar radiation duration, relative humidity). Results showed that all physico-chemical characteristics of Saanen goat milk varied significantly throughout the lactation period and years. The decrease of fat, protein, non-fat dry matter and lactose content in goat milk during the mid-lactation period was more pronounced than was previously reported in the literature. The highest values for these characteristics were recorded in the late lactation period. Observed variations were explained by negative correlation between THI and the physico-chemical characteristics of Saanen goat milk. This indicated that Saanen goats were very prone to heat stress, which implied the decrease of physico-chemical characteristics during hot summers.

In vitro and in vivo anti-tumor effects of selected platinum(IV) and dinuclear platinum(II) complexes against lung cancer cells.

In the present study, cytotoxic effects of cisplatin, the most usually used chemotherapeutic agent, were compared with new designed platinum(IV) ([PtCl4(en)] (en = ethylenediamine) and [PtCl4(dach)]) (dach = (±)-trans-1,2-diaminocyclohexane) and platinum(II) complexes ([{trans-Pt(NH3)2Cl}2(µ-pyrazine)](ClO4)2 (Pt1), [{trans-Pt(NH3)2Cl}2(µ-4,4'-bipyridyl)](ClO4)2DMF(Pt2),[{trans-Pt(NH3)2Cl}2(µ-1,2-bis(4pyridyl)ethane)](ClO4)2 (Pt3)), in vitro and in vivo against human and murine lung cancer cells, to determine anti-tumor potential of newly synthesized platinum-based drugs in the therapy of lung cancer. Results obtained by MTT [3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide], Lactate dehydrogenase and Annexin V/Propidium Iodide assays showed that, among all tested complexes, [PtCl4(en)] had the highest cytotoxicity against human and murine lung carcinoma cells in vitro. [PtCl4(en)] showed significantly higher cytotoxicity then cisplatin in all tested concentrations, mainly by inducing apoptosis in lung cancer cells. [PtCl4(en)] was well tolerated in vivo. Clinical signs of [PtCl4(en)]-induced toxicity, such as changes in food, water consumption or body weight, nephrotoxicity or hepatotoxicity was not observed in [PtCl4(en)]-treated mice. [PtCl4(en)] managed to increase presence of CD45+ leukocytes, including F4/80+ macrophages, CD11c+ dendritic cells, CD4+ helper and CD8+ cytotoxic T cells (CTLs) in the lungs, cytotoxic NK, NKT and CTLs in the spleens of tumor bearing mice, resulting with reduction of metastatic lesions in the lungs, indicating its potential to stimulate anti-tumor immune response in vivo. Due to its anti-tumor cytotoxicity, biocompatibility, and potential for stimulation of anti-tumor immune response, [PtCl4(en)] may be a good candidate for further testing in the field of medicinal chemistry.

Needle Terpenes as Chemotaxonomic Markers in Pinus: Subsections Pinus and Pinaster.

Chemical compositions of needle essential oils of 27 taxa from the section Pinus, including 20 and 7 taxa of the subsections Pinus and Pinaster, respectively, were compared in order to determine chemotaxonomic significance of terpenes at infrageneric level. According to analysis of variance, six out of 31 studied terpene characters were characterized by a high level of significance, indicating statistically significant difference between the examined subsections. Agglomerative hierarchical cluster analysis has shown separation of eight groups, where representatives of subsect. Pinaster were distributed within the first seven groups on the dendrogram together with P. nigra subsp. laricio and P. merkusii from the subsect. Pinus. On the other hand, the eighth group included the majority of the members of subsect. Pinus. Our findings, based on terpene characters, complement those obtained from morphological, biochemical, and molecular parameters studied over the past two decades. In addition, results presented in this article confirmed that terpenes are good markers at infrageneric level.

Chemotaxonomic Approach to the Central Balkan Sedum Species Based on Distribution of Triterpenoids in Their Epicuticular Waxes.

Triterpenoid distribution in epicuticular waxes of 20 central Balkan Sedum L. species and four out-groups of genera: Hylotelephium H. Ohba, Crassula L., Echeveria DC., and Kalanchoe Adans. were investigated for chemotaxonomic purposes. Identification and quantification of wax triterpenoids were performed by GC/MS and GC-FID analyses. Distribution of identified triterpenoids (oleanane, lupane, and taraxerane series), as a pattern in statistical analysis, indicated very good agreement with phylogeny and systematics, except members of series Rupestria Berger, in which case the distribution of triterpenoids did not support known classification in a satisfactory manner. According to the obtained clustering, Kalanchoe is shown as similar to Sedum samples, while the other out-groups were clearly separated.

Geographically Related Variation in Epicuticular Wax Traits of Pinus nigra Populations from Southern Carpathians and Central Balkans - Taxonomic Considerations.

The chemical composition of epicuticular waxes of nine populations from three Pinus nigra J. F. Arnold subspecies (namely subsp. nigra, subsp. banatica (Borbás) Novák, and subsp. pallasiana (Lamb.) Holmboe) from Southern Carpathians and central Balkan Peninsula were analyzed using GC/MS and GC/FID chromatography, and multivariate statistical techniques with respect to biogeography and taxonomy. In the needle waxes, four primary alcohols and 14 n-alkanes ranging from C21 to C33 were identified, and the most abundant compounds were the four odd-numbered n-alkanes C27 , C25 , C23 , and C29. Multivariate statistical analyses (CDA and CA) have shown existence of three P. nigra groups and suggested clinal differentiation as a mechanism of genetic variation across a geographic area: the first group consisted of the southernmost populations of subsp. pallasiana from Macedonia, the second consisted of the northernmost subsp. banatica populations from Romania, while all populations in Serbia described as three different subspecies (nigra, banatica, and pallasiana) formed the third group together with subsp. nigra population from Bosnia and Herzegovina. According to simple linear regression, geographic latitude and four bioclimatic parameters were moderately correlated with the contents of epicuticular wax compounds that are important in population discrimination, while stepwise multiple regression showed that latitude participated in most of the regression models for predicting the composition of the epicuticular waxes. These results agree with CDA and CA analysis, and confirmed the possibility of recognition of fine geographic differentiation of the analyzed P. nigra populations.

Distribution and variability of n-alkanes in epicuticular waxes of sedum species from the central Balkan Peninsula: chemotaxonomic importance.

For the first time, the n-alkane distribution and variability of the epicuticular waxes within 22 Sedum taxa was reported with focus on the chemotaxonomy of native Sedum representatives from the central Balkan Peninsula, compared to their relations with four other species of the Crassulaceae family. By GC/MS and GC-FID identification and quantification, it was established that n-alkanes C27 , C29 , C31 , C33 , and C35 were the dominant constituents of the examined epicuticular wax samples. Applying multivariate statistical analyses including agglomerative hierarchical clustering (AHC) and principal component analysis (PCA), the relation according to the n-alkane composition between the examined samples was established. It was shown that the n-alkane variability of the central Balkan Sedum species was considerable and that n-alkanes might not be very reliable taxonomic markers for these species.

The effect of heat treatment and skimming on precipitate formation in caprine and bovine milks.

Caprine and bovine milks have a similar overall gross composition, but vary considerably in the ratios of their casein components. These differences in colloidal casein micelles could affect directly or indirectly the heat stability of caprine and bovine milks at their natural pH. In the present work, the differences in colloidal stability of caprine and bovine milk have been studied by analysing the effect of heat treatment and skimming on precipitation of proteins. Raw and heated milk samples (70 °C/5 min, 80°C/5 min and 90°C/5 min) were centrifuged at 600, 2000, and 4500  g . The amount of precipitate formed after skimming was measured and the protein composition of both precipitates and supernatants analysed using the SDS-PAGE (sodium dodecyl sulphate polyacrylamide gel electrophoresis) and densitometry. In caprine milk, the heat treatment prior to skimming had a statistically significant effect on protein precipitation. Centrifugal force had a statistically significant effect on amount of precipitate for both milks, but the amount was 2 to 4 times higher for caprine milk. When defatting the milk for electrophoresis, a centrifugal force of 600  g appeared to be the most appropriate, in order to avoid protein loss and a possible error in the interpretation of results. Results of this study could also serve as the basis for further investigations on adjusting the skimming conditions for caprine milk in industrial dairy processing environment.

Invasive fungal disease in COVID-19 patients: a single-center prospective observational study.

Background: Invasive fungal diseases (IFDs) are caused by fungal infections that manifest as serious secondary infections in patients with COVID-19. The increased morbidity and mortality rates are most frequently observed in patients with COVID-19-associated pulmonary aspergillosis (CAPA) and COVID-19-associated candidiasis (CAC). CAPA is the most frequently encountered infection with an incidence rate of 0.7-7.7%, while CAC is a less common and less studied fungal infection in COVID-19 patients. Materials and methods: The present article is a prospective observational single-center study that was conducted between 1 September 2021 and 24 December 2021, involving 6,335 patients who were admitted to COVID Hospital "Batajnica," University Clinical Center of Serbia, Belgrade. Results: Of the 6,335 patients hospitalized during the four-month period of the study, 120 patients (1.86%) who had a proven diagnosis of IFD were included in the study. These patients were divided into two groups: CAPA patients (n = 63) and CAC patients (n = 56); however, one of the 120 patients was diagnosed with Cryptoccocus neoformans infection. The mean age of the study population was 65.7 ± 13.9 years, and 78 (65.5%) of them were men. The patients were identified to have the following non-malignant comorbidities: arterial hypertension in 62 (52.1%) patients, diabetes mellitus in 34 (28.65), pre-existing lung damage similar to that observed in COPD and asthma in 20 (16.8%), and chronic renal insufficiency in 13 (10.9%) patients. The hematological malignancies were found to be the most prevalent malignancies and were identified in 20 (16.8%) patients, particularly in CAPA patients [11 (17.5%); p < 0.041]. Fiberoptic bronchoscopy with bronchoalveolar lavage fluid (BALF) and microscopic examination confirmed the presence of fungal infections in 17 (14.3%) patients. Serology testing was also performed in the majority of cases. Antibodies against Aspergillus spp. and Candida spp. were predominantly found in CAPA patients (p < 0.001). The patients were also tested for the presence of (1-3)-ß-D glucan (p < 0.019), galactomannan, and mannan in the specimens. Blood cultures were found to be positive in 45 (37.8%) patients, mostly in CAC patients. Mechanical ventilation was applied in 41 (34.5%) patients, while a non-invasive technique, such as continuous positive airway pressure (CPAP) or high-flow nasal cannula (HFNC), was used in 20 (16.8%) patients. The following antifungals were administered: echinocandins in 42 (35.3%), voriconazole in 30 (25.2%), and fluconazole in 27 (22.7%) patients. Most of the patients received systemic corticosteroids (mainly methylprednisolone), while 11 (9.16%) received favipiravir, 32 (26.67%) remdesivir, 8 (6.67%) casirivimab/imdevimab, and 5 (4.16%) sotrovimab. The outcome was lethal in 76 (63.9%) patients, predominantly CAC patients (p < 0.001). Conclusion: Invasive fungal disease is a severe complication associated with COVID-19 and accounts for increased mortality in these patients. Early identification and appropriate treatment may provide a favorable outcome.

Incremental costs of hospital-acquired infections in COVID-19 patients in an adult intensive care unit of a tertiary hospital from a low-resource setting.

BACKGROUND: Hospital-acquired infections (HAIs) are a global public health problem and put patients at risk of complications, including death. HAIs increase treatment costs, but their financial impact on Serbia's healthcare system is unknown. Our goal was to assess incremental costs of HAIs in a tertiary care adult intensive care unit (ICU) that managed COVID-19 patients. METHODS: A retrospective study from March 6th to December 31st, 2020 included patients with microbiologically confirmed COVID-19 (positive rapid antigen test or real-time polymerase chain reaction) treated in the ICU of the Teaching Hospital for Infectious and Tropical Diseases, University Clinical Centre of Serbia. Demographic and HAI-specific data acquired in our ICU were collected, including total and stratified medical costs (services, materials, laboratory testing, medicines, occupancy costs). Median total and stratified costs were compared in relation to HAI acquisition. Linear regression modelling was used to assess incremental costs of HAIs, adjusted for age, biological sex, prior hospitalisation, Charlson Comorbidity Index (CCI), and Glasgow Coma Scale (GCS) on admission. Outcome variables were length of stay (LOS) in days and mortality. RESULTS: During the study period, 299 patients were treated for COVID-19, of which 214 were included. HAIs were diagnosed in 56 (26.2%) patients. Acinetobacter spp. was the main pathogen in respiratory (38, 45.8%) and bloodstream infections (35, 42.2%), the two main HAI types. Median total costs were significantly greater in patients with HAIs (€1650.4 vs. €4203.2, p < 0.001). Longer LOS (10.0 vs. 18.5 days, p < 0.001) and higher ICU mortality (51.3% vs. 89.3%, p < 0.001) were seen if HAIs were acquired. Patients with ≥ 2 HAIs had the highest median total costs compared to those without HAIs or with a single HAI (€1650.4 vs. €3343.4 vs. €7336.9, p < 0.001). Incremental costs in patients with 1 and ≥ 2 HAIs were €1837.8 (95% CI 1257.8-2417.7, p < 0.001) and €5142.5 (95% CI 4262.3-6022.7, p < 0.001), respectively. CONCLUSIONS: This is the first economic evaluation of HAIs in Serbia, showing significant additional costs to our healthcare system. HAIs prolong LOS and influence ICU mortality rates. Larger economic assessments are needed to enhance infection control practices.

Myositis-specific autoantibodies in a non-traveler, patient from a non-endemic country, with Plasmodium vivax malaria.

INTRODUCTION: Autoantibodies (AAb) are a hallmark of immune-mediated inflammatory diseases. Malaria is a parasitic disease caused by Plasmodium protozoa. Individuals with malaria may present with a wide range of symptoms. It is frequently linked to the development of different AAb. CASE DESCRIPTION: A 35-year-old male presented with repeated episodes of fever, malaise, myalgia, dark urine, and yellowish sclera. Initial diagnostic workup revealed severe Coombs-positive anemia, increased C-reactive protein, and procalcitonin, pathological liver tests, high concentration of serum IgE, IgG, IgM, IgA, positive antinuclear antibodies (ANA), and positive antineutrophil cytoplasmatic antibodies (ANCA). In addition, myositis-specific antibodies directed to polymiositis-scleroderma 75 protein (PmScl75), threonyl-tRNA synthetase (PL-7), alanyl-tRNA synthetase (PL-12), Mi-2 antigen (Mi-2), Ku DNA helicase complex (Ku), signal recognition particle (SRP), and antiaminoacyl tRNA synthetase (EJ) were detected. The patient was suspected of having systemic lupus erythematosus and sent to the Clinic of Allergy and Immunology for further evaluation and treatment. A peripheral blood film examined by the hematologist during an episode of fever revealed intra-erythrocytic parasitic forms of Plasmodium vivax (P. vivax). After being diagnosed with P. vivax malaria, he was transferred to the Clinic for Infective and Tropical Diseases. The therapy consisted of artesunate/mefloquine and prednisone led to a complete clinical recovery and autoantibodies gradually disappeared. CONCLUSIONS: Malaria would not normally be considered during the initial diagnostic workup in a non-traveler and a patient from a non-endemic country. However, a thorough parasitic evaluation in patients presenting with a broad range of autoantibodies might be of particular importance.