Association of the Brain-derived Neurotrophic Factor Val66Met Polymorphism with Body Mass Index, Fasting Glucose Levels and Lipid Status in Adolescents.

Brain-derived neurotrophic factor (BDNF) has an important role in energy balance. It suppresses food intake, reduces hepatic glucose production and converts white fat into brown fat in adipose tissue, leading to energy dissipation, lowered blood glucose and a lean phenotype. Studies have shown that the single nucleotide polymorphism (SNP) Val66Met within BDNF may be associated with obesity, insulin sensitivity, type 2 diabetes mellitus (T2DM) and dyslipidemia. The objective of the study was to investigate the association of the Val66Met polymorphism with body mass index (BMI), fasting glucose levels and lipid profile in Serbian adolescents. The study included 308 randomly selected healthy adolescents, 153 (49.68%) boys and 155 girls (50.32%), 15 years of age. Data including age, gender, height, weight, lipid profile and fasting glucose were recorded. Genotyping was performed by the polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) method. No association of this polymorphism was found with BMI and lipid profile. However, significant association was observed between this polymorphism and fasting blood glucose (FBG). Carriers of a Val/Val genotype had significantly higher mean values of fasting glucose level compared to carriers of Val/ Met and Met/Met genotypes (p = 0.01). To confirm these results multiple linear regression analysis was performed. Body mass index and gender were taken as covariates. Carriers of the Val/Val genotype had significantly higher levels of FBG (ß = -0.152, p = 0.02). A statistically significant association between BMI and glucose level was also observed (ß = 0.124,p = 0.033). This polymorphism could be associated with fasting glucose level in Serbian adolescents, thus further research would be of great interest to validate these results.

Exercise training in overweight and obese children: Recreational football and high-intensity interval training provide similar benefits to physical fitness.

This study compared the effects of recreational football and high-intensity interval training (HIIT) on body composition, muscular fitness, and cardiorespiratory fitness in overweight and obese children. Forty-two overweight/obese males aged 11-13 years [body mass index (BMI) >20.5 kg/m2 ] were randomly assigned to a recreational football training group (n = 14; 157.9 ± 5.8 cm; 63.7 ± 12.6 kg), HIIT group (n = 14; 163.8 ± 9.4 cm; 71.5 ± 10.5 kg), or nontraining control group (n = 14; 162.7 ± 9.3 cm; 67.4 ± 16.1 kg). Physical fitness components were measured at baseline and after 12 weeks of training at the same time of the day and under similar conditions, including body composition, muscular fitness (lower-body power, change-of-direction speed, and flexibility), and cardiovascular fitness (Yo-Yo Intermittent Endurance test distance, resting heart rate, and blood pressure). Lean body mass (4.3%, ES = 0.40; 95% CI: -0.48, 1.29; P = .382) and muscle mass 4.4% (ES = 0.40; 95% CI: -0.48, 1.29; P = .378) very likely increased in the recreational football group, while possible improvements were observed in the HIIT group (lean body mass: 2.5%, ES = 0.22; 95% CI: -0.62, 1.06; P = .607, muscle mass: 2.8%, ES = 0.23; 95% CI: -0.61, 1.07; P = .594). Only trivial increases were observed in the control group for lean body mass (0.5%, ES = 0.05; 95% CI: -0.70, 0.79; P = .906) and muscle mass (1.1%, ES = 0.09; 95% CI: -0.65, 0.83; P = .814). Significant differences were found between the recreational football and control groups in post-training body mass (P = .034) and body mass index (P = .017). Body fat very likely decreased in the recreational football group (-7.7%, ES = -0.41; 95% CI: -1.29, 0.48; P = .376) and possibly decreased in the HIIT group (-5.2%, ES = -0.22; 95% CI: -1.05, 0.62; P = .607), with a trivial reduction in the control group (-1.1%, ES = -0.04; 95% CI: -0.78, 0.70; P = .914). Very likely increases in lower-body power were evident in the recreational football (17.0%, ES = 0.76; 95% CI: -0.15, 1.66; P = .107) and control groups (16.1%, ES = 0.55; 95% CI: -0.20, 1.31; P = .156), while small improvements were observed in the HIIT group (6.0%, ES = 0.24; 95% CI: -0.60, 1.08; P = .580, possible). Likely to most likely improvements in Yo-Yo Intermittent Endurance test performance and change-of-direction speed were noted in the recreational football group (Yo-Yo: 79.8%, ES = 1.09; 95% CI: 0.16, 2.03; P = .025, change-of-direction speed: -10.6%, ES = -1.05; 95% CI: -1.98, -0.12; P = .031) and the HIIT group (Yo-Yo: 81.2%, ES = 1.03; 95% CI: 0.15, 1.92; P = .025, change-of-direction speed: -5.4%, ES = -0.91; 95% CI: -1.79, -0.04; P = .045). Diastolic blood pressure likely decreased in the recreational football (-8.6%, ES = -0.74; 95% CI: -1.64, 0.17; P = .116) and HIIT groups (-9.8%, ES = -0.57; 95% CI: -1.40, 0.30; P = .195), with a possible increase in the control group (1.2%, ES = 0.21; 95% CI: -0.53, 0.96; P = .068). Recreational football and HIIT elicited improvements in all muscular and cardiorespiratory fitness measures. In contrast, the control group, which performed only physical education classes, increased body mass, BMI, and fat mass. Therefore, additional activities such as recreational football or HIIT might counter the prevalence of overweight and obesity in children.

Implementation of a Clinical Decision Support Tool for Stool Cultures and Parasitological Studies in Hospitalized Patients.

There is substantial evidence that stool culture and parasitological examinations are of minimal to no value after 3 days of hospitalization. We implemented and studied the impact of a clinical decision support tool (CDST) to decrease the number of unnecessary stool cultures (STCUL), ova/parasite (O&P) examinations, and Giardia/Cryptosporidium enzyme immunoassay screens (GC-EIA) performed for patients hospitalized >3 days. We studied the frequency of stool studies ordered before or on day 3 and after day 3 of hospitalization (i.e., categorical orders/total number of orders) before and after this intervention and denoted the numbers and types of microorganisms detected within those time frames. This intervention, which corresponded to a custom-programmed hard-stop alert tool in the Epic hospital information system, allowed providers to override the intervention by calling the laboratory, if testing was deemed medically necessary. Comparative statistics were employed to determine significance, and cost savings were estimated based on our internal costs. Before the intervention, 129/670 (19.25%) O&P examinations, 47/204 (23.04%) GC-EIA, and 249/1,229 (20.26%) STCUL were ordered after 3 days of hospitalization. After the intervention, 46/521 (8.83%) O&P examinations, 27/157 (17.20%) GC-EIA, and 106/1,028 (10.31%) STCUL were ordered after 3 days of hospitalization. The proportions of reductions in the number of tests performed after 3 days and the associated P values were 54.1% for O&P examinations (P < 0.0001), 22.58% for GC-EIA (P = 0.2807), and 49.1% for STCUL (P < 0.0001). This was estimated to have resulted in $8,108.84 of cost savings. The electronic CDST resulted in a substantial reduction in the number of evaluations of stool cultures and the number of parasitological examinations for patients hospitalized for more than 3 days and in a cost savings while retaining the ability of the clinician to obtain these tests if clinically indicated.

Bone mineral density at different sites and vertebral fractures in Serbian postmenopausal women.

OBJECTIVES: This randomized study aimed to evaluate the correlation between bone mineral densities (BMD) measured at different sites and the frequency of vertebral fractures in a group of Serbian postmenopausal women. METHOD: BMD was measured in 130 naïve postmenopausal women by dual X-ray absorptiometry (DXA) at the ultra-distal part of the forearms, at the hip and at the lumbar spine. At each of the measurement sites, the patients were categorized as osteoporotic, or osteopenic, or in the reference range. Vertebral fractures were examined using thoracic and lumbar spine radiography. RESULTS: A T-score at different skeletal sites showed discordance in the site-specific region. Vertebral fractures were found in 58.82% of patients with hip osteopenia, in 45% with forearm osteopenia and in 54.54% with lumbar spine osteoporosis. CONCLUSIONS: The study confirmed that the reduction of BMD depends on age and choice of measurement site. The best correlation was obtained in the women with osteopenia at all measurement sites. The discovery of vertebral fractures by lateral thoracic and lumbar spine radiography improves prompt treatment. Reference values of BMD do not exclude vertebral fractures. Of vertebral fractures, 72.5% were asymptomatic and thus spine radiographies are obligatory. Currently discussed is the position of DXA for measuring BMD as a method of detection for patients at risk of fracture.

Evaluation of functional outcome measured by modified Rankin scale in rtPA treated patients with acute ischemic stroke.

Aim of our study was to assess functional outcome measured by modified Rankin scale (mRS) in patients that were treated with thrombolytic therapy-recombinant tissue plasminogen activator (rtPA) after acute ischemic stroke. The study included 100 participants that were treated after acute ischemic stroke. Analyzed parameters included: gender; age groups: age 54 and below (Groupup to-54), 55-64 (Group55-64), 65-74 (Group65-74), and 75 and above (Group75-up); cerebral blood flow (CBF) and cerebral blood volume (CBV). Considering time of rtPA administration, we analyzed 3 groups: between 1-2 hours from stroke onset (Time1-2h), 2-3 hours (Time2-3h) and 3-4.5 hours (Time3h-up). NIHSS scores were analyzed: NIHSS 1-at admission and NIHSS 2-at discharge from hospital; and mRS values: RANKIN 1-at admission and RANKIN 2-at discharge from hospital. There is significant reduction in NIHSS and mRS scores between two measurements for all groups of evaluated parameters. CBF, CBV and NIHSS values at admission significantly correlated with mRS scores at admission (p<0.01), as well as with mRS scores at discharge except for CBF where statistical significance was (p=0.019). Significantly lower values of NIHSS at admission (p<0.01), CBF values (p<0.01) and CBV values (p<0.01) are noticed in the group with mRS≤2. Early induction of rtPA treatment in patients with acute ischemic stroke within first 4.5 hours significantly increases positive treatment outcome in both genders and for all evaluated age groups. Favorable outcome (mRS≤2) at the time of discharge from hospital is significantly associated with lower NIHSS values at admission.

First Report of Fusarium Root Rot of Stored Carrot Caused by Fusarium avenaceum in Serbia.

Carrot (Daucus carota L. subsp. sativus (Hoffm.) Thell., Apiaceae), a widely consumed antioxidant-rich plant, is among the major vegetable crops grown in Serbia, with average annual production of 65,400 tons on approximately 7,000 ha (4). In May 2013, a severe root rot was observed on approximately 20% of cold-stored carrot roots originating from Gospodinci, South Backa District, Serbia. Symptoms included dry rot of the collar and crown as well as large, brown to dark brown, circular, sunken lesions on the stored roots. Frequently, abundant whitish mycelium was observed covering the surface of the colonized roots. To determine the causal agent, small pieces of infected tissue were surface-disinfested with 2% NaOCl without rinsing, air-dried, and placed on potato dextrose agar. Five single-spore isolates obtained from collar and crown tissue sections, as well as nine isolates from root sections, all formed abundant, cottony white to pale salmon fungal colonies with reddish orange pigment on the reverse surface of the agar medium when grown at 25°C under 12 h of fluorescent light per day. All recovered isolates formed numerous, three- to six-septate, hyaline, needle-like, straight to slightly curved, fusoid macroconidia (30 to 80 × 4 to 5.5 µm, average 58.3 × 4.9 µm, n = 100 spores) each with a tapering apical cell. Microconidia of all isolates were generally scarce, two- to four-septate, spindle-shaped, and 15 to 35 × 3 to 5 µm (average 21.3 × 4.2 µm). Chlamydospores were not observed. Based on these morphological characteristics, the pathogen was identified as Fusarium avenaceum (Fries) Saccardo (1). The pathogenicity on carrot was tested for isolate 19-14 by inoculating each of five carrot roots surface-disinfected with 2% NaOCl, by placing a mycelial plug into the surface of a wound created with a cork borer. Carrot roots inoculated with sterilized PDA plugs served as a negative control treatment. After 5 days of incubating the roots at 25°C, root rot symptoms identical to those observed on the source carrot plants developed on all inoculated roots, and the pathogen was re-isolated from each of these roots using the same procedure descibed above. There were no symptoms on the control roots. Morphological species identification was confirmed by sequencing the translation elongation factor (EF-1α) gene (2). Total DNA was extracted directly from fungal mycelium of isolate 19-14 with a DNeasy Plant Mini Kit (Qiagen, Hilden, Germany), and PCR amplification was performed with primer pair EF-1/EF-2 (2). Sequence analysis of the EF-1α gene revealed 100% nucleotide identity of isolate 19-14 (GenBank Accession No. KM102536) with the EF-1α sequences of two F. avenaceum isolates from Canada (KC999504 from rye and JX397864 from Triticum durum). To our knowledge, this is the first report of F. avenaceum causing collar, crown, and root rots of stored carrot in Serbia. Since F. avenaceum can produce several mycotoxins, including moniliformin, acuminatopyrone, and chrysogine (3), the presence of this pathogen on stored carrots could represent a significant constraint for carrot production in Serbia, for both direct yield losses and potential mycotoxin contamination. References: (1) J. F. Leslie and B. A. Summerell. The Fusarium Laboratory Manual, Blackwell Publishing, London, UK, 2006. (2) K. O'Donnell et al. Proc. Natl. Acad. Sci. U.S.A. 95:2044, 1998. (3) J. L. Sorenson. J. Agric. Food Chem. 57:1632, 2009. (4) Statistical Office, Republic of Serbia. Retrieved from http://webrzs.stat.gov.rs in May 2014.

First Report of Cucumber mosaic virus in Tulipa sp. in Serbia.

Tulips (Tulipa sp. L.), popular spring-blooming perennials in the Liliaceae family, are one of the most important ornamental bulbous plants, which have been cultivated for cut flower, potted plant, garden plant, and for landscaping. In May 2013, during a survey to determine the presence of Cucumber mosaic virus (CMV, Cucumovirus, Bromoviridae) on ornamentals in Serbia, virus-like symptoms, including the presence of bright streaks, stripe and distortion of leaves, and reduced growth and flower size, were observed in an open field tulip production in the Krnjaca locality (a district of Belgrade, Serbia). Disease incidence was estimated at 20%. Symptomatic tulip plants were collected and tested for the presence of CMV by double-antibody sandwich (DAS)-ELISA using commercial diagnostic kit (Bioreba, AG, Reinach, Switzerland). Commercial positive and negative controls were included in each ELISA. Of the six tulip plants tested, all were positive for CMV. In bioassay, five plants of each Chenopodium quinoa, Nicotiana tabacum 'Samsun,' and N. glutinosa were mechanically inoculated with sap from selected ELISA-positive sample (79-13) using 0.01 M phosphate buffer (pH 7). Chlorotic local lesions on C. quinoa, and severe mosaic and leaf malformations on N. tabacum 'Samsun' and N. glutinosa, were observed 5 and 14 days post-inoculation, respectively. All mechanically inoculated plants were positive for CMV in DAS-ELISA testing. For further confirmation of CMV presence in tulip, total RNAs from all ELISA-positive symptomatic tulip plants were extracted with the RNeasy Plant Mini Kit (Qiagen, Hilden, Germany). Reverse transcription (RT)-PCR was performed with the One-Step RT-PCR Kit (Qiagen) using specific primer pair CMVCPfwd and CMVCPrev (1), which flank conserved fragment of the RNA3 including the entire coat protein (CP) gene and part of 3'- and 5'-UTRs. Total RNAs obtained from the Serbian watermelon CMV isolate (GenBank Accession No. JX280942) and healthy tulip leaves served as the positive and negative controls, respectively. The RT-PCR products of 871 bp were obtained from all six samples that were serologically positive to CMV, as well as from the positive control. No amplicon was recorded in the healthy control. The amplified product which derived from isolate 79-13 was purified (QIAquick PCR Purification Kit, Qiagen), directly sequenced in both directions using the same primer pair as in RT-PCR, deposited in GenBank (KJ854451), and analyzed by MEGA5 software (4). Sequence comparison of the complete CP gene (657 nt) revealed that the Serbian isolate 79-13 shared the highest nucleotide identity of 99.2% (99% amino acid identity) with CMV isolates from Japan (AB006813) and the United States (S70105). To our knowledge, this is the first report on the occurrence of CMV causing mosaic on Tulipa sp. in Serbia. Taking into account vegetative reproduction of tulips and the large scale of international trade with tulip seeding material, as well as wide host range of CMV including a variety of ornamentals (2,3), this is a very important discovery representing a serious threat for the floriculture industry in Serbia. References: (1) K. Milojevic et al. Plant Dis. 96:1706, 2012. (2) M. Samuitiene and M. Navalinskiene. Zemdirbyste-Agriculture 95:135, 2008. (3) D. Sochacki. J. Hortic. Res. 21:5, 2013. (4) K. Tamura et al. Mol. Biol. Evol. 28:2731, 2011.

First Report of Fusarium Wilt of Strawberry Caused by Fusarium oxysporum in Serbia.

Strawberry (Fragaria × ananassa Duch.) is the third most important berry crop in Serbia with average production ranging from 30,000 to 35,000 t on approximately 5,000 ha (2). In June 2013, symptoms of wilt and whole plant collapse were observed on approximately 25% plants growing in commercial strawberry crop of cv. Alba in the locality of Zablace (Moravica district). Initial symptoms included leaf chlorosis and wilt, followed by withering and necrosis of older leaves and reduced fruit production, eventually leading to plant collapse and desiccations. Internal vascular tissues of the crown showed distinct brown reddish discoloration. Three small pieces of infected roots, petioles, or crown vascular tissues were surface disinfested with 2% NaOCl and placed on five potato dextrose agars (PDA) per sample. After 7 days incubation at 23°C under 12 h of fluorescent light, nine monoconidial isolates were obtained (1) forming colonies with light purple mycelia. Colonies produced numerous hyaline, oval to ellipsoid microconidia (5 to 15 × 2.5 to 4.5 µm, average 8.45 × 2.25 µm), 3 to 5 septate fusoid macroconidia with pedicellate bases (20 to 50 × 2.70 to 6 µm, average 32.35 × 3.25 µm from 100 measured) and chlamydospores. Morphological and growth features were similar to the descriptions of Fusarium oxysporum Schlechtend emend. Snyder & Hansen (1). Pathogenicity of one selected isolate (97-13) was tested by dipping for 15 min the roots of five plants of each cultivar: Alba, Arosa, Clery, and Roxana into a conidial suspension (1 × 106 conidia/ml) harvested from a 7-day-old culture on PDA. Control plants were dipped in sterile distilled water. The inoculated plants were transplanted into pots containing sterilized peat and maintained in the greenhouse at 25°C. Thirty to thirty-five days post-inoculation, all plants developed wilt symptoms and vascular discoloration of crown tissues from which F. oxysporum was successfully re-isolated using the same method as for isolation. No symptoms were observed on any of the control plants. Morphological identification was confirmed by amplification and sequencing of a portion of the translation elongation factor-1 alpha (EF-1α) gene. Total DNA was extracted directly from fungal mycelium with a DNeasy Plant Mini Kit (Qiagen, Hilden, Germany) and PCR amplification performed with primers EF-1/EF-2 (4). Sequence analysis of EF-1α region revealed that Serbian isolate 97-13 (GenBank Accession No. KJ647280) shared 99 to 100% identity with the F. oxysporum sequences in GenBank. To our knowledge, this is the first report of Fusarium wilt on strawberry in Serbia. The presence of a new and potentially harmful disease may represent a serious constraint for strawberry production in Serbia. References: (1) J. F. Leslie and B. A. Summerell. The Fusarium Laboratory Manual, Blackwell Publishing, London, UK, 2006. (2) M. Nikolic et al. Acta Hort. 842:615, 2009. (3) K. O'Donnell et al. Proc. Natl. Acad. Sci. USA 95:2044, 1998.

Effects of Candida on insulin secretion of human adult pancreatic islets and possible onset of diabetes.

This study aims to determine the origin of Candida contamination of pancreatic tissue cultures, as well as its influence on insulin secretory activity of the pancreatic islets. Pancreatic tissue was obtained after pancreatectomy in patients who had chronic pancreatitis or benign tumours. Islets were isolated under aseptic conditions by a manual method. Microbiological analysis was performed by standard procedures and secretory activity was determined on the first, third and seventh day of cultivation. Insulin stimulation index (SI) on the first day of incubation was 0.665 +/- 0.082 and 0.982 +/- 0.167 for sterile and infected cultures, respectively (expressed as means +/- SE). On the third day of cultivation, the SI for sterile cultures was 0.645 +/- 0.071 while these value were higher in contaminated cultures (1.252 +/- 0.413). On the seventh day, SI was 0.853 +/- 0.032 and 1.239 +/- 0.169 for sterile and infected cultures, respectively (P = 0.05). Analysis of results for the first, third and seventh day of incubation and comparison of both groups showed that SI was 0.721 +/- 0.041 for sterile cultures, while for contaminated cultures it was higher by 37.68% (SI = 1.157 +/- 0.154; P = 0.01). The results show that cell culture contamination originates from an original pancreatic tissue infection, and that Candida can provoke an elevated level of insulin secretion in such patients, thus increasing chances for the onset of diabetes.

Effect of low temperature cultivation on insulin secretory of human pancreatic islets.

The experiment compared the physiological function (insulin secretory capacity) and membrane integrity of human adult pancreatic islets incubated in culture at 37°C and 24°C. Pancreatic tissue was digested with Collagenase XI, using a non-automated method. Cultures were incubated at 37°C and 24°C. Secretory capacity of the islets is determined by measuring of the stimulation index (SI) on the 1st, 3rd and 7th day of cultivation. Membrane integrity of the islets was determined by dithizone staining. Both groups of examined cultures show a slight increase in SI during the incubation. However islets incubated at 24°C show higher SI values than those incubated at 37°C on the 1st, 3rd and 7th day of incubation. And on the first day of incubation, this difference was statistically significant (p <0.05). Islets incubated at 37°C showed preservation of membrane integrity, the islets are regular spherical shape, while those incubated at 24°C lose such an organization. During the seven-day cultivation, islets incubated at a standard temperature of 37°C show less preserve physiological functions in relation to cultures incubated at 24°C, but islets incubated at 37°C show more regular morphological forms.

First Report of Tomato spotted wilt virus on Chrysanthemum in Serbia.

In July 2011, greenhouse-grown chrysanthemum hybrid plants (Chrysanthemum × morifolium) with symptoms resembling those associated with tospoviruses were observed in the Kupusina locality (West Backa District, Serbia). Disease incidence was estimated at 40%. Symptomatic plants with chlorotic ring spots and line patterns were sampled and tested by double antibody sandwich (DAS)-ELISA using polyclonal antisera (Bioreba AG, Reinach, Switzerland) against the two of the most devastating tospoviruses in the greenhouse floriculture industry: Tomato spotted wilt virus (TSWV) and Impatiens necrotic spot virus (INSV) (2). Commercial positive and negative controls and extracts from healthy chrysanthemum tissue were included in each ELISA. TSWV was detected serologically in 16 of 20 chrysanthemum samples and all tested samples were negative for INSV. The virus was mechanically transmitted from ELISA-positive chrysanthemum samples to five plants each of both Petunia × hybrida and Nicotiana tabacum 'Samsun' using chilled 0.01 M phosphate buffer (pH 7) containing 0.1% sodium sulfite. Inoculated plants produced local necrotic spots and systemic chlorotic/necrotic concentric rings, consistent with symptoms caused by TSWV (1). The presence of TSWV in ELISA-positive chrysanthemum plants and N. tabacum'Samsun' was further confirmed by conventional reverse transcription (RT)-PCR. Total RNAs were extracted with an RNeasy Plant Mini Kit (Qiagen, Hilden, Germany). RT-PCR was performed with the One-Step RT-PCR Kit (Qiagen) using primers TSWVCP-f/TSWVCP-r specific to the nucleocapsid protein (N) gene (4). A Serbian isolate of TSWV from tobacco (GenBank Accession No. GQ373173) and RNA extracted from a healthy chrysanthemum plant were used as positive and negative controls, respectively. An amplicon of the correct predicted size (738-bp) was obtained from each of the plants assayed, and that derived from chrysanthemum isolate 529-11 was purified (QIAqick PCR Purification Kit, Qiagen) and sequenced (JQ692106). Sequence analysis of the partial N gene, conducted with MEGA5 software, revealed the highest nucleotide identity of 99.6% (99% amino acid identity) with 12 TSWV isolates deposited in GenBank originating from different hosts from Italy (HQ830186-87, DQ431237-38, DQ398945), Montenegro (GU355939-40, GU339506, GU339508), France (FR693055-56), and the Czech Republic (AJ296599). The consensus maximum parsimony tree obtained on a 705-bp partial N gene sequence of TSWV isolates available in GenBank revealed that Serbian TSWV isolate 529-11 from chrysanthemum was clustered in the European subpopulation 2, while the Serbian isolates from tomato (GU369723) and tobacco (GQ373172-73 and GQ355467) were clustered in the European subpopulation 1 denoted previously (3). The distribution of TSWV in commercial chrysanthemum crops is wide (2). To our knowledge, this is the first report of TSWV infecting chrysanthemum in Serbia. Since chrysanthemum popularity and returns have been rising rapidly, the presence of TSWV may significantly reduce quality of crops in Serbia. References: (1) Anonymous. OEPP/EPPO Bull. 34:271, 2004. (2) Daughtrey et al. Plant Dis. 81:1220, 1997. (3) I. Stankovic et al. Acta Virol. 55:337, 2011. (4) A. Vucurovic et al. Eur. J. Plant Pathol. 133:935, 2012.

First Report of Tomato spotted wilt virus on Brugmansia sp. in Serbia.

Brugmansia (Brugmansia spp.), also known as Angel's trumpet, is a perennial shrub in the Solanaceae that is a popular landscape plant in the tropics and subtropics, and potted plant in temperate regions. In April 2012, virus-like symptoms including chlorotic leaf patterns and curling followed by necrosis and distortion of leaves were observed on five outdoor-grown brugmansia plants in a private garden in Mackovac, Rasina District, Serbia. Symptomatic leaves were tested for the presence of several common ornamental viruses including Tomato spotted wilt virus (TSWV), Impatiens necrotic spot virus (INSV), Cucumber mosaic virus (CMV), and Tobacco mosaic virus (TMV) by commercial double-antibody sandwich (DAS)-ELISA diagnostic kits (Bioreba AG, Reinach, Switzerland). Commercial positive and negative controls and extract from healthy brugmansia leaves were included in each ELISA. TSWV was detected serologically in all five brugmansia samples and all tested samples were negative for INSV, CMV, and TMV. The virus was mechanically transmitted from an ELISA-positive sample (41-12) to five plants of each Petuina × hybrida and Nicotiana glutinosa. Inoculated P. × hybrida plants showed local necrotic lesions and N. glutinosa showed mosaic and systemic necrosis 4 and 12 days post-inoculation, respectively, which were consistent with symptoms caused by TSWV (1). For further confirmation of TSWV infection, reverse transcription (RT)-PCR was performed with the OneStep RT-PCR (Qiagen, Hilden, Germany) using a set of TSWV-specific primers, TSWV CP-f and TSWV CP-r (4), designed to amplify a 738-bp fragment of the nucleocapsid protein (N) gene. Total RNAs from naturally infected brugmansia and symptomatic N. glutinosa plants were extracted using the RNeasy Plant Mini Kit (Qiagen). Total RNAs obtained from the Serbian tobacco isolate of TSWV (GenBank Accession No. GQ373173) and healthy brugmansia plants were used as positive and negative controls, respectively. The expected size of the RT-PCR product was amplified from symptomatic brugmansia and N. glutinosa but not from healthy tissues. The amplified product derived from the isolate 41-12 was sequenced directly after purification with the QIAquick PCR Purification kit (Qiagen), deposited in GenBank (JX468080), and subjected to sequence analysis by MEGA5 software (3). Sequence comparisons revealed that the Serbian isolate 41-12 shared the highest nucleotide identity of 99.9% (99.5% amino acid identity) with an Italian TSWV isolate P105/2006RB (DQ915946) originating from pepper. To our knowledge, this is the first report of TSWV on brugmansia in Serbia. Due to the increasing popularity and economic importance of brugmansia as an ornamental crop, thorough inspections and subsequent testing for TSWV and other viruses are needed. This high-value ornamental plant may act also as reservoir for the virus that can infect other ornamentals and cultivated crops, considering that TSWV has a very broad host range (2). References: (1) Anonymous. OEPP/EPPO Bull. 34:271, 2004. (2) G. Parrella et al. J. Plant Pathol. 85:227, 2003. (3) K. Tamura et al. Mol. Biol. Evol. 28:2731, 2011. (4) A. Vucurovic et al. Eur. J. Plant Pathol. 133:935, 2012.

Abdominal aortic aneurysm volume and relative intraluminal thrombus volume might be auxiliary predictors of rupture-an observational cross-sectional study.

Objectives: The study aimed to identify differences and compare anatomical and biomechanical features between elective and ruptured abdominal aortic aneurysms (AAAs). Methods: Data (clinical, anatomical, and biomechanical) of 98 patients with AAA, 75 (76.53%) asymptomatic (Group aAAA) and 23 (23.46%) ruptured AAA (Group rAAA), were prospectively collected and analyzed. Anatomical, morphological, and biomechanical imaging markers like peak wall stress (PWS) and rupture risk equivalent diameter (RRED), comorbid conditions, and demographics were compared between the groups. Biomechanical features were assessed by analysis of Digital Imaging and Communication in Medicine images by A4clinics (Vascops), and anatomical features were assessed by 3Surgery (Trimensio). Binary and multiple logistic regression analysis were used and adjusted for confounders. Accuracy was assessed using receiving operative characteristic (ROC) curve analysis. Results: In a multivariable model, including gender and age as confounder variables, maximal aneurysm diameter [MAD, odds ratio (OR) = 1.063], relative intraluminal thrombus (rILT, OR = 1.039), and total aneurysm volume (TAV, OR = 1.006) continued to be significant predictors of AAA rupture with PWS (OR = 1.010) and RRED (OR = 1.031). Area under the ROC curve values and correct classification (cc) for the same parameters and the model that combines MAD, TAV, and rILT were measured: MAD (0.790, cc = 75%), PWS (0.713, cc = 73%), RRED (0.717, cc = 55%), TAV (0.756, cc = 79%), rILT (0.656, cc = 60%), and MAD + TAV + rILT (0.797, cc = 82%). Conclusion: Based on our results, in addition to MAD, other important predictors of rupture that might be used during aneurysm surveillance are TAV and rILT. Biomechanical parameters (PWS, RRED) as valuable predictors should be assessed in prospective clinical trials. Similar studies on AAA smaller than 55 mm in diameter, even difficult to organize, would be of even greater clinical value.

The Role of Systemic Oxidative Status in Coronary Arterial and Peripheral Venous Blood of Patients with Unstable Angina Pectoris.

(1) Background: We aimed to analyze the oxidative status of patients with unstable angina pectoris (UA), as well as to determine the correlation of these parameters between coronary arterial and peripheral venous blood samples. (2) Methods: The study included 47 human subjects with UA and 45 control subjects. We performed clinical examinations, hemodynamic and coronary angiography measures. Also, in the blood samples, we measured routine laboratory markers and the concentration of pro-oxidants: index of lipid peroxidation (TBARS), superoxide anion radical (O2-), hydrogen peroxide (H2O2) and nitrites (NO2-), while antioxidant parameters were determined from red blood cells: reduced glutathione (GSH), catalase (CAT) and superoxide dismutase (SOD). All parameters were determined spectrophotometrically. (3) Results: Significantly higher values of TBARS and all measured antioxidants SOD, CAT and GSH were observed in the coronary arterial blood of the UA group relative to coronary arterial blood of the control subjects. On the other hand, in the peripheral venous blood samples, a significantly lower GSH value was found in the UA group compared to the control. (4) Conclusions: This study has shown that the majority of changes in all measured redox markers are found in coronary blood, especially related to the activity of antioxidant components. In patients with an unstable form of angina, prooxidants (superoxide anion radical and index of lipid peroxidation) and endogenous antioxidants (catalase, superoxide dismutase and reduced glutathione) are in direct correlation with the course of ischemic disease. Future studies, where participants would be randomized depending on symptom duration, are necessary to confirm these conclusions.

First Report of Tomato spotted wilt virus Infecting Onion and Garlic in Serbia.

In June 2011, extensive bleaching and numerous small whitish spots on leaves were observed in an onion (Allium cepa) seed crop as well as chlorotic spots and streaks in the neighboring garlic (A. sativum) bulb crop in the Aleksandrovo locality (Central Banat District, Serbia). Affected plants occurred throughout the field and disease incidence was estimated at 60% in the onion and 40% in the garlic crop. A high population of Thrips tabaci that was found in both crops, and local necrotic spots on Petunia × hybrida mechanically inoculated with infected onion or garlic sap by a chilled 0.01 M phosphate buffer, pH 7.0, containing 0.1% sodium sulfite (1), suggested the presence of a Tospovirus. For these reasons, sampled symptomatic onion and garlic plants were tested for the presence of Tomato spotted wilt virus (TSWV) and Iris yellow spot virus (IYSV) using commercial double-antibody sandwich-ELISA diagnostic kits (Bioreba AG, Reinach, Switzerland). Commercial positive and negative controls and extracts from healthy onion and garlic tissue were included in each ELISA. Of the 18 onion and 10 garlic plants tested, 16 and 7 samples, respectively, were positive for TSWV, and all were negative for IYSV. The identity of TSWV was further confirmed by conventional reverse transcription (RT)-PCR analysis. Total RNAs were extracted with an RNeasy Plant Mini Kit (Qiagen, Hilden, Germany) and RT-PCR was performed with the One-Step RT-PCR Kit (Qiagen) using TSWV-specific forward (5'-GGTTAAGCTCACTAAGAAARCA-3') and reverse primers (5'-TTTAACYCCRAACATTTCATAGA-3'), designed to amplify a 738-bp fragment of the nucleocapsid protein (N) gene. Total RNAs obtained from plants infected with a Serbian isolate of TSWV (GenBank Accession No. GQ373173) and healthy onion garlic plants were used as positive and negative controls, respectively. An amplicon of the expected size was produced from the 16 onion and 7 garlic ELISA-positive plants, but not from healthy controls. The amplified products derived from the two selected isolates, 114-11 from onion and 115-11 from garlic, were sequenced directly after purification with the QIAquick PCR Purification kit (Qiagen); the sequences obtained were allocated GenBank Accession Nos. JQ619234 and JQ619235, respectively. Sequence analysis of the partial N gene, conducted with MEGA5 software (4), revealed 99.9% nucleotide identity (100% amino acid identity) between the two Serbian Allium isolates. Serbian onion and garlic isolates showed the highest nucleotide identities of 100% and 99.9% with Serbian summer squash isolate (JF303081) and tobacco isolate from Montenegro (GU369729), respectively. Well-established in many European countries, TSWV has been reported as an important constraint to the production of tomato, pepper, tobacco, and ornamentals (2), but the information on TSWV naturally infecting Allium spp. is limited. The presence of TSWV on onion and garlic in Serbia revealed that its known host range has expanded in Europe. To our knowledge, other than Marchoux's unpublished data (3), there are no other reports of garlic as a natural host of TSWV. The TSWV presence on Allium spp. represents a serious threat for these crops in Serbia, considering that it is prevalent in other crops in the area and its vectors are widespread. References: (1) Anonymous. OEPP/EPPO Bull. 34:271, 2004. (2) H. R. Pappu et al. Virus Res. 141:219, 2009. (3) G. Parrella et al. J. Plant Pathol. 85:227, 2003. (4) K. Tamura et al. Mol. Biol. Evol. 28:2731, 2011.

First Report of Cucumber mosaic virus Infecting Watermelon in Serbia.

In June 2012, field-grown watermelon plants (Citrullus lanatus L.) with virus-like symptoms were observed in Silbas locality, South Backa District of Serbia. Plants infected early in the growing season showed severe symptoms including stunting, mosaic, mottling, blistering, and leaf curling with reduced leaf size, while those infected at later stages exhibited only a mild mosaic. Affected plants were spread across the field and disease incidence was estimated at 40%. Thirteen symptomatic watermelon plants were sampled and analyzed by double-antibody sandwich (DAS)-ELISA using a commercial diagnostic kit (Bioreba AG, Reinach, Switzerland) against the most important watermelon viruses: Cucumber mosaic virus (CMV), Watermelon mosaic virus (WMV), Zucchini yellow mosaic virus (ZYMV), Papaya ringspot virus (PRSV), and Squash mosaic virus (SqMV) (1). Commercial positive and negative controls and an extract from healthy watermelon tissue were included in each ELISA. Serological analyses showed that all plants were positive for CMV and negative for ZYMV, WMV, PRSV, and SqMV. The virus was mechanically transmitted from an ELISA-positive sample (449-12) to five plants of each Citrullus lanatus 'Creamson sweet' and Chenopodium amaranticolor using 0.01 M phosphate buffer (pH 7) with Serbian CMV isolate from Cucurbita pepo 'Olinka' (GenBank Accession No. HM065510) and healthy watermelon plants as positive and negative controls, respectively. Small necrotic lesions on C. amaranticolor and mild mosaic with dark green vein banding on watermelon leaves were observed on all inoculated plants 5 and 14 days post-inoculation, respectively. For further confirmation of CMV infection, reverse transcription (RT)-PCR was performed with the One-Step RT-PCR Kit (Qiagen, Hilden, Germany) using specific primers CMVCPfwd (5'-TGCTTCTCCRCGARWTTGCGT-3') and CMVCPrev (5'-CGTAGCTGGATGGACAACCCG-3'), designed to amplify an 871-bp fragment of the RNA3 including the whole CP gene. Total RNA from 12 naturally infected and five mechanically infected watermelon plants was extracted with the RNease Plant Mini Kit (Qiagen). Total RNA obtained from the Serbian CMV isolate (HM065510) and healthy watermelon plants were used as positive and negative controls, respectively. The expected size of RT-PCR products were amplified from all naturally and mechanically infected watermelon plants but not from healthy tissues. The PCR product derived from isolate 449-12 was purified and directly sequenced using the same primer pair as in RT-PCR (JX280942) and analyzed by MEGA5 software (3). Sequence comparison of the complete CP gene (657 nt) revealed that the Serbian isolate 449-12 shared the highest nucleotide identity of 98.9% (99.1% amino acid identity) with the Spanish melon isolate (AJ829777) and Syrian tomato isolate (AB448696). To our knowledge, this is the first report of CMV on watermelon in Serbia. CMV is widely distributed within the Mediterranean basin where it has a substantial impact on many agricultural crops (2) and is often found to be prevalent during pumpkin and squash surveys in Serbia (4). The presence of CMV on watermelon could therefore represent a serious threat to this valuable crop in Serbia. References: (1) L. M. da Silveira et al. Trop. Plant Pathol. 34:123, 2009. (2) M. Jacquemond. Adv. Virus Res. 84:439, 2012. (3) K. Tamura et al. Mol. Biol. Evol. 28:2731, 2011. (4) A. Vucurovic et al. Eur. J. Plant Pathol. 133:935, 2012.

First Report of Zucchini yellow mosaic virus in Watermelon in Serbia.

During a survey of cucurbit viruses in the Gornji Tavankut locality (North Backa District), Serbia in June 2011, field-grown (a surface of 1.8 ha) watermelon plants (Citrullus lanatus [Thunb.] Matsum and Nakai) with mild mosaic symptoms were observed. Large numbers of Aphis gossypii were colonizing the crop. A total of 26 samples, six from plants exhibiting mosaic and 20 from asymptomatic plants, were analyzed by double-antibody sandwich-ELISA using polyclonal antisera virus (Bioreba AG, Reinach, Switzerland) against three cucurbit-infecting viruses known to infect Cucurbita pepo in Serbia: Zucchini yellow mosaic virus (ZYMV), Cucumber mosaic virus, and Watermelon mosaic virus (3). Commercial positive and negative controls were included in ELISA analysis. Only six symptomatic samples tested positive for ZYMV, but no other tested viruses were found. The virus was mechanically transmitted from a representative ELISA-positive watermelon sample (550-11) to five plants of C. pepo 'Ezra F1' and severe mosaic was noticed 10 days after inoculation. For further confirmation of ZYMV infection, total RNA from a naturally infected watermelon plant and symptomatic C. pepo 'Ezra F1' plants were extracted with the RNeasy Plant Mini Kit (Qiagen, Hilden, Germany). Reverse transcription (RT)-PCR was performed with the One-Step RT-PCR Kit (Qiagen) using primer pair ZY-2 and ZY-3 (2). Total RNA obtained from a Serbian isolate of ZYMV from pumpkin (GenBank Accession No. HM072432) and healthy watermelon plants were used as positive and negative controls, respectively. The expected sizes of the RT-PCR products (1,186 bp) were amplified from naturally and mechanically infected symptomatic samples, but not from healthy tissues. The amplified product that derived from isolate 550-11 was purified (QIAquick PCR Purification Kit, Qiagen), sequenced in both directions, deposited in GenBank (Accession No. JN561294), and subjected to sequence analysis using MEGA4 software. Sequence comparisons revealed a high nucleotide identity of 99.9 to 99.8% and 100 to 99.6% amino acid identity for the CP gene with Serbian ZYMV isolates from C. pepo (Accession Nos. JF308188, HM072431, and HM072432). The nucleotide and deduced amino acid sequences of the entire CP gene (837 nt) of the Serbian ZYMV isolate from watermelon shared 99.9 to 93.7% and 100 to 96.8% identity, respectively, with innumerous isolates of ZYMV deposited in the GenBank (e.g., Accession Nos. AJ420012-17 and FJ705262). To our knowledge, this is the first report of ZYMV spreading its host range to watermelon in Serbia. ZYMV infection has been responsible for severe epidemics on cucurbits throughout the world (1). The presence of ZYMV on watermelon could therefore represent a serious threat for this valuable crop in Serbia, especially considering that it is prevalent in other cucurbit crops in the country and the vectors are widespread. References: (1) H. Lecoq et al. Virus Res. 141:190, 2009. (2) K. G. Thomson et al. J. Virol. Methods 55:83, 1995. (3) A. Vucurovic et al. Pestic. Phytomed. (Belgrade) 24:85, 2009.

The importance of timing in surgical treatment of unruptured symptomatic aneurysm of abdominal aorta.

AIM OF STUDY: Aim of this study is to define an entity of unruptured symptomatic AAA, to examine the influence of timing of the surgical treatment and to analyze the results of the treatment of unruptured symptomatic AAA in acute expansion. MATERIALS AND METHOD: The study is designed as retrospective analysis of 390 operatively treated patients in the last five years at the Clinics of Vascular Surgery in Novi Sad. All patients were grouped into four categories: elective operative surgical treatment, surgical treatment 24 hours after the admission through the Department of Urgent Surgery with an urgent CT diagnosis (in first 2 hours), surgical treatment within 24 hours since the admission through the Department of Urgent Surgery with an urgent CT diagnosis (in first 2 hours) and immediate surgical treatment of ruptured AAA. RESULTS: In the period from Jan 1, 2005 to Dec 31, 2009, 390 patients with AAA were operatively treated. 89 patients had ruptured AAA, 52 were operated 24 hours after the urgent admission, 18 patients were operated in the first 24 hours after the urgent admission and 231 patients were planned for elective surgery. Mortality rates between the groups were as follows: elective surgery-5.1 %, patients operated 24 hours after the urgent admission 7.2 %, patients operated in the first 24 hours after the urgent admission 23 %, and patients who had ruptured AAA 34 %. CONCLUSION: Considering the obtained data, it can be concluded that the treatment of unruptured symptomatic AAA is related to a higher risk of postoperative mortality in relation to an elective surgery. Moreover, surgical treatment in the first 24 hours after the urgent admission of unruptured symptomatic AAA has higher rate of mortality and morbidity compared to surgical treatment 24 hours after the urgent admission of the patients, so we can conclude that the early (semi) elective surgery is a method of choice for the treatment of unruptured symptomatic AAA in acute expansion (Tab. 2, Fig. 2, Ref. 21).

siMS score- method for quantification of metabolic syndrome, confirms co-founding factors of metabolic syndrome.

Background: Adipose tissue is a dynamic endocrine organ, a highly active metabolic tissue, and an important source of cytokines. Inflammatory factors play an important role in visceral obesity associated with insulin resistance (IR), metabolic syndrome (MS), hypertension, non-alcoholic fatty liver disease (NAFLD), diabetes mellitus type 2 (DM2), endothelial dysfunction (ED) and atherosclerosis. Objectives: To examine corelation of siMS score, as a quantification method for metabolic syndrome (MS), with insulin resistance, glucoregulation parameters, as with other co-founding factors of MS, inflammation and thrombosis factors, microalbuminuria, uric acid, fatty liver index (FLI) and homocysteine. Methods: The study included 451 obese individuals with pre-metabolic syndrome (pre-MS) and MS (age 16-75, body mass index (BMI) > 25kg/m2) classified into two groups: I-age 10-30 (167 patients); II-age 31-75 (284 patients). International Diabetes Federation (IDF) classification was applied for diagnosing metabolic syndrome. Patients with less than three criteria indicated below were considered pre-metabolic syndrome. siMS risk score was used. Results: siMS score increased with age: I-3.03 ± 0.87, II-3.27 ± 0.90. siMS score correlated with associated factors of MS: hyperinsulinemia and IR, ALT, gama-GT, FLI, uric acid in both groups and CRP (p < 0.01) in group I. Correlations in II group: siMS score with PAI-1 (p = 0.01), microalbuminuria (p = 0.006), homocysteine ​​(p = 0.076). Conclusion: Correlation of siMS score with HOMA-IR confirmed that hyperinsulinism and insulin resistance are in the basis of MS. Correlation of siMS score with parameters of NAFLD, CRP, PAI-1, uric acid, microalbuminuria and homocysteine indicates that they are significant co-founding factors of MS. Correlation of siMS score with PAI-1, microalbuminuria, homocysteine, indicates higher risk for progression of endothelial dysfunction and atherosclerosis with age.