The relationship between polymorphism of four serotonic genes (<Emphasis Type="Italic">5-HTTL, 5-HT1A, 5-HT2A</Emphasis>, and <Emphasis Type="Italic">MAOA</Emphasis>) and personality traits in wrestlers and control group

The molecular genetic analysis of serotonin system genes (5-HTTL, 5-HT1A, 5-HT2A, and MAOA) in male and female wrestlers and in a control group has been performed. Comparative population genetics analysis of the 5-HTTL gene showed the highest frequency of the SS genotype 5-HTTLPR in athletes (p = 0.04), as well as a trend to higher frequencies of united genotypes of the locus 5-HTTLPR VNTR and SNP rs25531—S _A S _A (p = 0.06) compared to the control group. As for the markers of 5-HT1A (rs6295), 5-HT2A (rs6311), and MAOA (VNTR), we found no significant differences between the groups tested. Using an NEO PI-R questionnaire, possible correlations between the genotypes and psychological traits were analyzed in the studied samples. It was demonstrated that the athletic achievements of upper-tier athletes are associated with their lower openness to experience and higher conscientiousness. It was revealed that the factors of gender and 5-HT1A genotype should influence the self-rating for openness to experience. Furthermore, a combined effect of the level of sports achievements and 5-HT2A genotypes was found, as well as gender and 5-HT1A genotypes, on the self-rating for conscientiousness.     

Mammary candidiasis: molecular-based detection of <Emphasis Type="Italic">Candida</Emphasis> species in human milk samples

In this prospective and monocentric study, we investigated the performance of a commercialized real-time polymerase chain reaction (RT-PCR) test system for the specific detection of DNA from Candida albicans, C. dubliniensis, C. glabrata, C. krusei, C. lusitaniae, C. parapsilosis, and C. tropicalis in human milk samples of patients suspicious of mammary candidiasis. For this purpose, 43 breast-feeding women with characteristic symptoms of mammary candidiasis and 40 asymptomatic controls were enrolled. By culture, Candida spp. were detected in 8.8 % (4/46) and 9.3 % (4/43) of patient and control samples, respectively. Candida albicans (2/46), C. parapsilosis (1/46), and C. guilliermondii (1/46) were present in patient samples, and C. lusitaniae (3/43) and C. guilliermondii (1/43) were present in the controls. After RT-PCR was applied, Candida spp. were found to be present in 67.4 % (31/46) and 79.1 % (34/43) of patient and control samples investigated, respectively. PCR detection of C. albicans and C. parapsilosis revealed only a low sensitivity and specificity of 67.4 % and 41.9 %, respectively. Our data do not support the use of Candida RT-PCR for sensitive and specific diagnosis of mammary candidiasis.     

Synthesis of two potential NK<Subscript>1</Subscript>-receptor ligands using [1-<Superscript>11</Superscript>C]ethyl iodide and [1-<Superscript>11</Superscript>C]propyl iodide and initial PET-imaging

Background

The previously validated NK₁-receptor ligand [O-methyl-¹¹C]GR205171 binds with a high affinity to the NK₁-receptor and displays a slow dissociation from the receptor. Hence, it cannot be used in vivo for detecting concentration changes in substance P, the endogenous ligand for the NK₁-receptor. A radioligand used for monitoring these changes has to enable displacement by the endogenous ligand and thus bind reversibly to the receptor. Small changes in the structure of a receptor ligand can lead to changes in binding characteristics and also in the ability to penetrate the blood-brain barrier. The aim of this study was to use carbon-11 labelled ethyl and propyl iodide with high specific radioactivity in the synthesis of two new and potentially reversible NK₁-receptor ligands with chemical structures based on [O-methyl-¹¹C]GR205171.

Methods

[1-¹¹C]Ethyl and [1-¹¹C]propyl iodide with specific radioactivities of 90 GBq/μmol and 270 GBq/μmol, respectively, were used in the synthesis of [O-methyl-¹¹C]GR205171 analogues by alkylation of O-desmethyl GR205171. The brain uptake of the obtained (2S,3S)-N-(1-(2- [1-¹¹C]ethoxy-5-(3-(trifluoromethyl)-4H-1,2,4-triazol-4-yl)phenyl)ethyl)-2-phenylpiperidin-3-amine (I) and (2S,3S)-2-phenyl-N-(1-(2- [1-¹¹C]propoxy-5-(3-(trifluoromethyl)-4H-1,2,4-triazol-4-yl)phenyl)ethyl)piperidin-3-amine (II) was studied with PET in guinea pigs and rhesus monkeys and compared to the uptake of [O-methyl-¹¹C]GR205171.

Results

All ligands had similar uptake distribution in the guinea pig brain. The PET-studies in rhesus monkeys showed that (II) had no specific binding in striatum. Ligand (I) had moderate specific binding compared to the [O-methyl-¹¹C]GR205171. The ethyl analogue (I) displayed reversible binding characteristics contrary to the slow dissociation rate shown by [O-methyl-¹¹C]GR205171.

Conclusion

The propyl-analogue (II) cannot be used for detecting changes in NK₁-ligand levels, while further studies should be performed with the ethyl-analogue (I).

     

Mutations in Hotspot Regions of <Emphasis Type="Italic">ERG11</Emphasis> Gene in Fluconazole Resistant Isolates of <Emphasis Type="Italic">Candida albicans</Emphasis> in Guilan Province,Northern Iran

Background

Widespread use of azoles has resulted in rapid development of azole resistance in Candida albicans strains. Mutations in ERG11, a target enzyme of azoles, alter the binding ability of azoles to this enzyme and result in the development of resistant strains. In this study, we evaluated ERG11 mutations in fluconazole resistant isolates of C. albicans.

Materials and methods

In this study, 60 clinical samples were isolated from Guilan hospitals. Then differential tests were used to identify C. albicans strains. Disc diffusion and MIC tests were used to the analyze fluconazole susceptibility. Then, the resistant isolates were evaluated by PCR and sequencing methods for ERG11 mutations.

Results

Of 60 clinical samples, 40 C. albicans strains were identified through specific symptoms. Susceptibility tests showed that four C. albicans strains were resistant to high dose fluconazole (≥512 μg/mL). In all resistant isolates was found missense mutations such as K291N, C470G and Q474R and three isolates had premature nonsense mutation (Y477stop).

Discussion

Our study indicates that the level of fluconazole resistance in C. albicans strains is high in Guilan province and other drugs should be used in resistant infections. It seems that missense mutations in four isolates play role in azole resistance. However in three isolates premature stop codon may be involved in high dose resistance. And it is suggested that in fourth isolates another mechanisms introduce increase of resistance dose in combination with missense mutation in ERG11. Results of this study suggest that in patients by high dose of resistance do not use azole because of mutations that decrease azole effects.

     

Change of annexin binding of monocytes as an expression of cellular response to <Emphasis Type="Italic">Candida albicans</Emphasis>: down-regulation in severe sepsis

To study the differences of monocyte activation by albicans and non-albicans species of Candida and its change in sepsis, peripheral blood mononuclear cells were isolated from 17 healthy volunteers and 26 patients with severe sepsis/shock, and incubated in the absence/presence of heat-killed (HK) isolates of four different Candida species and purified β-D-glucan from C.albicans. Experiments were repeated in the presence and absence of inhibitors of intracellular activation pathways. Expression of annexin V on cells membranes of monocytes and lymphocytes, cytoplasmic activity of caspase-3, and DNA fragmentation of monocytes were studied. Membrane expression of annexin V on viable monocytes of healthy volunteers decreased significantly after incubation with C.albicans but not with non-albicans species. The decrease was dose-dependent from the Candida inoculum and by the concentration of β-D-glucan. A relationship with inhibition of apoptosis was found as the activity of caspase-3 activity, and the level of DNA fragmentation were also decreased. Incubation in the absence/presence of inhibitors showed that the decrease by annexin V expression resulted by activation of the dectin-1 pathway and Raf-1 by β-D glucan. The decrease of annexin V(+)/PI(?) expression was not shown on monocytes of patients with severe sepsis/shock, where no effect of inhibitors was found. Decrease of annexin V binding on monocytes can be viewed as a selective response to C.albicans partly effected through activation of dectin-1. This response is down-regulated after a septic insult.     

Spread and exchange of <Emphasis Type="Italic">bla</Emphasis><Subscript>NDM-1</Subscript> in hospitalized neonates: role of mobilizable genetic elements

To investigate the mobilizable elements associated with bla _NDM-1 in Enterobacteriaceae isolated from septicaemic neonates at a NICU in India, during December, 2008–2011. An attempt was also made to understand whether there was a pattern in the temporal acquisition of bla _NDM-1 within the unit. Transferability of carbapenem resistance was tested by conjugation and transformation. Plasmid types and addiction systems were analysed. The genetic background of bla _NDM-1 and association with class 1 integron were evaluated by PCR mapping. RFLP was carried out to discriminate plasmids of same incompatibility group. Transfer of carbapenem resistance was successful in 13/15 cases. bla _NDM-1 was associated with different plasmid scaffolds (IncFII, IncL/M, IncN, IncR, IncHIB-M/FIB-M), IncF type being the prevalent one. Addiction systems ccdAB and hok/sok were associated with transferable plasmids. Genetic structures surrounding bla _NDM-1 showed its association with at least a remnant of ISAba125 at its 5′-end. The spread of NDM-1 was not related to class 1 integron which possessed resistance determinants against trimethoprim (dfrA12, dfrA1, dfrA5), streptomycin (aadA2, aacA4), and rifampicin (arr-3). RFLP showed that three isolates possessed the same FII/FIIs plasmid; two of these three isolates were from a single neonate, implying interspecies transfer of bla _NDM-1. The predominance of FII plasmids and ISAba125 along with bla _NDM-1 was noted, but no specific pattern in the temporal acquisition of mobile genetic elements could be identified. To the best of our knowledge, this report is the first to inform the in-vivo interspecies plasmid transfer event of bla _NDM-1 in a neonate.     

Effect of aqueous extract of <Emphasis Type="Italic">Tinospora cordifolia</Emphasis> on functions of peritoneal macrophages isolated from CCl<Subscript>4</Subscript> intoxicated male albino mice

Background

The current practice of ingesting phytochemicals for supporting the immune system or fighting infections is based on centuries-old tradition. Macrophages are involved at all the stages of an immune response. The present study focuses on the immunostimulant properties of Tinospora cordifolia extract that are exerted on circulating macrophages isolated from CCl₄ (0.5 ml/kg body weight) intoxicated male albino mice.

Methods

Apart from damaging the liver system, carbon tetrachloride also inhibits macrophage functions thus, creating an immunocompromised state, as is evident from the present study. Such cell functions include cell morphology, adhesion property, phagocytosis, enzyme release (myeloperoxidase or MPO), nitric oxide (NO) release, intracellular survival of ingested bacteria and DNA fragmentation in peritoneal macrophages isolated from these immunocompromised mice. T. cordifolia extract was tested for acute toxicity at the given dose (150 mg/kg body weight) by lactate dehydrogenase (LDH) assay.

Results

The number of morphologically altered macrophages was increased in mice exposed to CCl₄. Administration of CCl₄ (i.p.) also reduced the phagocytosis, cell adhesion, MPO release, NO release properties of circulating macrophages of mice. The DNA fragmentation of peritoneal macrophages was observed to be higher in CCl₄ intoxicated mice. The bacterial killing capacity of peritoneal macrophages was also adversely affected by CCl_4. However oral administration of aqueous fraction of Tinospora cordifolia stem parts at a dose of 40 mg/kg body weight (in vivo) in CCl₄ exposed mice ameliorated the effect of CCl₄, as the percentage of morphologically altered macrophages, phagocytosis activity, cell adhesion, MPO release, NO release, DNA fragmentation and intracellular killing capacity of CCl₄ intoxicated peritoneal macrophages came closer to those of the control group. No acute toxicity was identified in oral administration of the aqueous extract of Tinospora cordifolia at a dose of 150 mg/kg body weight.

Conclusion

From our findings it can be suggested that, polar fractions of Tinospora cordifolia stem parts contain major bioactive compounds, which directly act on peritoneal macrophages and have been found to boost the non-specific host defenses of the immune system. However, the molecular mechanism of this activity of Tinospora cordifolia on immune functions needs to be elucidated.

     

Epidemiology and reporting of candidaemia in Belgium: a multi-centre study

The primary aim of this study was to collect national epidemiological data on candidaemia and to determine the reporting time of species identification and antifungal susceptibility in clinical practice. During a 1-year period (March 2013 until February 2014), every first Candida isolate from each episode of candidaemia was included prospectively from 30 Belgian hospitals. Identification and susceptibility testing were performed according to local procedures and isolates were sent to the National Reference Center for Mycosis. Species identification was checked by matrix-assisted laser desorption/ionisation time-of-flight mass spectrometry (MALDI-TOF MS) and internal transcribed spacer (ITS) sequencing in case no reliable identification was obtained by MALDI-TOF MS. Antifungal susceptibility testing was performed according to the European Committee on Antimicrobial Susceptibility Testing (EUCAST) methodology. A total of 355 isolates were retrieved from 338 patients. The mean incidence rate of candidaemia was 0.44 (range: 0.07 to 1.43) per 1000 admissions or 0.65 (range: 0.11 to 2.00) per 10,000 patient days. Candida albicans was most frequently found (50.4 %), followed by C. glabrata (27.3 %) and C. parapsilosis sensu lato (9.8 %). The overall resistance to fluconazole was 7.6 %, ranging from 3.9 % in C. albicans to 20.0 % in C. tropicalis. Only one C. glabrata isolate was resistant to the echinocandins. Four days after blood culture positivity, 99.7 % of the identifications and 90.3 % of the antifungal profiles were reported to the treating clinician. Candidaemia incidence rates differed up to 20-fold among Belgian hospitals; no clear factors explaining this difference were identified. The overall antifungal resistance rates were low but high azole resistance rates were recorded in C. tropicalis.     

The <Emphasis Type="Italic">NLRP3</Emphasis> and <Emphasis Type="Italic">CASP1</Emphasis> gene polymorphisms are associated with developing of acute coronary syndrome: a case-control study

The protein products of NLRP3 and CASP1 genes are involved in the cleavage of pro-IL-1B and pro-IL-18 leading to the active cytokines, which play an important role in the development of the acute coronary syndrome (ACS). The aim of the present study was to evaluate whether NLRP3 and CASP1 gene polymorphisms are biomarkers of ACS susceptibility in Mexican population. Two polymorphisms of the CASP1 gene [G+7/in6A (rs501192) and A10370-G Exon-6 (rs580253)] and one of the NLRP3 gene [UTR′3 G37562-C (rs10754558)] were genotyped by 5′ exonuclease TaqMan assays in a group of 617 patients with ACS and 609 control individuals. Under recessive model, the CASP1 G+7/in6A polymorphism was associated with an increased risk of developing ACS when compared to healthy controls (OR = 1.76, 95% CI 1.08–2.86, P _Res  = 0.022). In the same way, under recessive model, the CASP1 A10370-G was associated with increased risk of ACS (OR = 1.75, 95% CI 1.07–2.85, P _Res  = 0.025). Moreover, under co-dominant, dominant, over-dominant, and additive models, the NLRP3 UTR′3 G37562-C was associated with a decreased risk of ACS (OR = 0.45, 95%CI 0.22–0.92, P _Co-dom  = 0.006; OR = 0.61, 95%CI 0.44–0.84, P _Dom  = 0.002; OR = 0.67, 95%CI 0.48–0.94, P _Over-dom  = 0.02; and OR = 0.65, 95%CI 0.50–0.94, P _Add  = 0.02, respectively). In summary, this study demonstrates that the G+7/in6A and A10370-G polymorphisms of the CASP1 gene are associated with increased risk of developing ACS, whereas the UTR′3 G37562-C polymorphism of the NLRP3 gene is associated with a decreased risk of developing ACS in Mexican population.     

From medicinal plant extracts to defined chemical compounds targeting the histamine H<Subscript>4</Subscript> receptor: <Emphasis Type="Italic">Curcuma longa</Emphasis> in the treatment of inflammation

Objectives

The aim was to evaluate the activity of seven medicinal, anti-inflammatory plants at the hH₄R with focus on defined chemical compounds from Curcuma longa.

Materials

Activities were analyzed with membrane preparations from Sf9 cells, transiently expressing the hH₄R, G_αi2 and G_β1γ2 subunits.

Methods

From the methanolic extract of C. longa curcumin (1), demethoxycurcumin (2) and bis(4-hydroxy-cinnamoyl)methane (3) were isolated, purified with HPLC (elution-time 10.20, 9.66, 9.20 min, respectively) and together with six additional extracts, were characterized via radioligand binding studies at the hH₄R.

Results

Compounds from C. longa were the most potent ligands at the hH₄R. They exhibited estimated K _i values of 4.26–6.26 µM (1.57–2.31 µg/mL) (1); 6.66––8.97 µM (2.26–3.04 µg/mL) (2) and 10.24–14.57 µM (3.16–4.49 µg/mL) (3) (95% CI). The estimated K _i value of the crude extract of curcuma was 0.50–0.81 µg/mL. Fractionated curcumin and the crude extract surpassed the effect of pure curcumin with a K _i value of 5.54 µM or 2.04 µg/mL [95% CI (4.47–6.86 µM), (1.65–2.53 µg/mL)].

Conclusion

Within this study, defined compounds of C. longa were recognized as potential ligands and reasonable lead structures at the hH₄R. The mode of anti-inflammatory action of curcumin was further elucidated and the role of extracts in traditional phytomedicine was strengthened.

     

Expression of <Emphasis Type="Italic">Clonorchis sinensis</Emphasis> GIIIsPLA<Subscript>2</Subscript> protein in baculovirus-infected insect cells and its overexpression facilitating epithelial-mesenchymal transition in Huh7 cells via AKT pathway

Although prior studies confirmed that group III secretory phospholipase A₂ of Clonorchis sinensis (CsGIIIsPLA₂) had stimulating effect on liver fibrosis by binding to LX-2 cells, large-scale expression of recombinant protein and its function in the progression of hepatoma are worth exploring. Because of high productivity and low lipopolysaccharides (LPS) in the Sf9-baculovirus expression system, we firstly used this system to express the coding region of CsGIIIsPLA₂. The molecular weight of recombinant CsGIIIsPLA₂ protein was about 34 kDa. Further investigation showed that most of the recombinant protein presented intracellular expression in Sf9 insect cell nucleus and could be detected only into cell debris, which made the protein purification and further functional study difficult. Therefore, to study the role of CsGIIIsPLA₂ in hepatocellular carcinoma (HCC) progression, CsGIIIsPLA₂ overexpression Huh7 cell model was applied. Cell proliferation, migration, and the expression level of epithelial-mesenchymal transition (EMT)-related molecules (E-cadherin, N-cadherin, α-catenin, Vimentin, p300, Snail, and Slug) along with possible mechanism were measured. The results indicated that CsGIIIsPLA₂ overexpression not only inhibited cell proliferation and promoted migration and EMT but also enhanced the phosphorylation of AKT in HCC cells. In conclusion, this study supported that CsGIIIsPLA₂ overexpression suppressed cell proliferation and induced EMT through the AKT pathway.     

Initial antifungal strategy does not correlate with mortality in patients with candidemia

The incidence of Candida bloodstream infections (BSIs) has increased over time, especially in medical wards. The objective of this study was to evaluate the impact of different antifungal treatment strategies on 30-day mortality in patients with Candida BSI not admitted to intensive care units (ICUs) at disease onset. This prospective, monocentric, cohort study was conducted at an 1100-bed university hospital in Rome, Italy, where an infectious disease consultation team was implemented. All cases of Candida BSIs observed in adult patients from November 2012 to April 2014 were included. Patients were grouped according to the initial antifungal strategy: fluconazole, echinocandin, or liposomal amphotericin B. Cox regression analysis was used to identify risk factors significantly associated with 15-day and 30-day mortality. During the study period, 130 patients with candidemia were observed (58 % with C. albicans, 7 % with C. glabrata, and 23 % with C. parapsilosis). The first antifungal drug was fluconazole for 40 % of patients, echinocandin for 57.0 %, and liposomal amphotericin B for 4 %. During follow-up, 33 % of patients died. The cumulative mortality 30 days after the candidemia episode was 30.8 % and was similar among groups. In the Cox regression analysis, clinical presentation was the only independent factor associated with 15-day mortality, and Acute Physiology and Chronic Health Evaluation (APACHE) II score and clinical presentation were the independent factors associated with 30-day mortality. No differences in 15-day and 30-day mortality were observed between patients with and without C. albicans candidemia. In patients with candidemia admitted to medical or surgical wards, clinical severity but not the initial antifungal strategy were significantly correlated with mortality.     

Antibody to <Emphasis Type="Italic">Chlamydia trachomatis</Emphasis> proteins,TroA and HtrA,as a biomarker for <Emphasis Type="Italic">Chlamydia trachomatis</Emphasis> infection

We studied whether antibody to two chlamydial proteins (TroA and HtrA) could be used as biomarkers of Chlamydia trachomatis infection. Methods: Recombinant proteins C. trachomatis TroA and HtrA were used as antigens in enzyme immunoassay (EIA). Both IgG and IgA antibody responses were studied. Results: IgG or IgA antibody to either protein was infrequently detected in sera from healthy blood donors or virgin girls. Patients attending the STI Clinic and patients with perihepatitis had often IgG antibody against TroA (25 and 50 % respectively) and HtrA (21 and 38 % respectively). Especially in sera from patients with chlamydial perihepatitis, the A_450nm values with TroA were high (mean 1.591). A positive correlation between C. trachomatis MIF antibody and TroA (r? =?0.7) as well as HtrA (r? = 0.5) antibody was observed in sera from STI clinic patients and perihepatitis patients. Individuals with C. trachomatis infection and positive serology already when seeking medical attention had higher A_450nm values for TroA (0.638) and HtrA (0.836) than patients with no marker of previous exposure or with no infection (0.208 and 0.234 respectively). Diagnosis of genital C. trachomatis infection is often NAAT-based, whereas serology has little value in testing for uncomplicated genital C. trachomatis infection. TroA and HtrA antibodies are potential biomarkers for evaluation of ascending and repeated C. trachomatis infection.     

Development of <Emphasis Type="Italic">T</Emphasis><Subscript><Emphasis Type="Italic">m</Emphasis></Subscript>-shift genotyping method for detection of cat-derived <Emphasis Type="Italic">Giardia lamblia</Emphasis>

To develop T _m -shift genotyping method for detection of cat-derived Giardia lamblia, two sets of primers with two GC-rich tails of unequal length attached to their 5′-end were designed according to two SNPs (BG434 and BG170) of β-giardin (bg) gene, and specific PCR products were identified by inspection of a melting curve on real-time PCR thermocycler. A series of experiments on the stability, sensitivity, and accuracy of T _m -shift method was tested, and clinical samples were also detected. The results showed that two sets of primers based on SNP could distinguish accurately between assemblages A and F. Coefficient of variation of T _m values of assemblage A and F was 0.14 and 0.07% in BG434 and 0.10 and 0.11% in BG170, respectively. The lowest detection concentration was 4.52 × 10^?5 and 4.88 × 10^?5 ng/μL samples of assemblage A and F standard plasmids. The T _m -shift genotyping results of ten DNA samples from the cat-derived G. lamblia were consistent with their known genotypes. The detection rate of clinical samples by T _m -shift was higher than that by microscopy, and their genotyping results were in complete accordance with sequencing results. It is concluded that the T _m -shift genotyping method is rapid, specific, and sensitive and may provide a new technological mean for molecular detection and epidemiological investigation of the cat-derived G. lamblia.     

<Emphasis Type="Italic">CDKN1A</Emphasis> histone acetylation and gene expression relationship in gastric adenocarcinomas

CDKN1A is a tumor suppressor gene involved in gastric carcinogenesis and is a potential target for histone deacetylase inhibitor-based therapies. Upregulation of CDKN1A is generally observed in several cell lines after histone deacetylase inhibitor treatment; however, little is known about the histone acetylation status associated with this gene in clinical samples, including gastric tumor tissue samples. Therefore, our goal was to quantify the H3K9 and H4K16 acetylation levels associated with three CDKN1A regions in 21 matched pairs of gastric adenocarcinoma and corresponding adjacent non-tumor samples by chromatin immunoprecipitation and to correlate these data with the gene expression. Our results demonstrated that the ?402, ?20, and +182 CDKN1A regions showed a significantly increased acetylation level in at least one of the histones evaluated (p < 0.05, for all comparisons), and these levels were positively correlated in gastric tumors. However, an inverse correlation was detected between both H3K9 and H4K16 acetylation at the ?402 CDKN1A region and mRNA levels in gastric tumors (r = ?0.51, p = 0.02; r = ?0.60, p < 0.01, respectively). Furthermore, increased H4K16 acetylation at the ?20 CDKN1A region was associated with gastric tumors of patients without lymph node metastasis (p = 0.04). These results highlight the complexity of these processes in gastric adenocarcinoma and contribute to a better understanding of CDKN1A regulation in carcinogenesis.     

Dissemination of <Emphasis Type="Italic">bla</Emphasis><Subscript>OXA-370</Subscript> is mediated by IncX plasmids and the Tn6435 transposon

In Enterobacteriaceae, the bla_OXA-48-like genes have been identified on plasmids in different regions of the world. The OXA-370 is a plasmid-encoded OXA-_48-like enzyme reported in two distinct regions of Brazil. Recently, we demonstrate that the bla_OXA-370 gene is disseminated among several Enterobacteriaceae species and clones, indicating a high potential for dissemination. In this work, we described for the first time the complete nucleotide sequence of six plasmids harboring the bla_OXA-370 gene. Complete DNA sequencing using the Illumina platform and annotation of the plasmids showed that they belonged to incompatibility groups IncX and had in average 70 kbp. The bla_OXA-370 gene is located in a composite transposon containing four genes encoding transposases, named Tn6435. In this study, highly similar plasmids were detected in different Enterobacteriaceae genera.     

Molecular characterisation and antifungal susceptibility of clinical <Emphasis Type="Italic">Cryptococcus deuterogattii</Emphasis> (AFLP6/VGII) isolates from Southern Brazil

Cryptococcosis, caused by Cryptococcus gattii sensu lato, is an emerging disease that was initially found in (sub)tropical regions but recently expanded to temperate regions. Cryptococcus gattii s.l. infections are mostly encountered in healthy individuals, frequently affecting both lungs and the central nervous system (CNS). Usually, C. gattii s.l. is less susceptible to antifungal compounds than its counterpart, C. neoformans s.l. We studied 18 clinical C. gattii s.l. isolates with amplified fragment length polymorphism (AFLP) fingerprinting, mating-typing, multi-locus sequence typing (MLST) and antifungal susceptibility testing. All isolates were C. deuterogattii (genotype AFLP6/VGII), 14 were mating-type α and four were type a. Amphotericin B, itraconazole, voriconazole, posaconazole and isavuconazole showed high activity, with minimum inhibitory concentration (MIC) ranges of 0.063–0.25, 0.031–0.25, 0.031–0.25, 0.031–0.25 and <0.016–0.25 μg mL^?1, respectively. Fluconazole and flucytosine had high geometric mean MICs of 2.07 and 3.7 μg mL^?1, respectively. Most cases occurred in immunocompetent patients (n?=?10; 55.6 %) and CNS involvement was the most common clinical presentation (n?=?14; 77.8 %). Three patients (16.7 %) showed sequelae, hyperreflexia, dysarthria, diadochokinesia, anosmia and upper limb weakness. In conclusion, all infections were caused by C. deuterogattii (AFLP6/VGII) and the majority of patients were immunocompetent, with the CNS as the most affected site. All antifungal drugs had high in vitro activity against C. deuterogattii isolates, except fluconazole and flucytosine.     

Presence of a functional (TTAGG)<Subscript><Emphasis Type="Italic">n</Emphasis></Subscript> telomere-telomerase system in aphids

The structure of the telomeres of four aphid species (Acyrthosiphon pisum, Megoura viciae, Myzus persicae and Rhopalosiphum padi) was evaluated by Southern blotting and fluorescent in situ hybridization, revealing that each chromosomal end consists of a (TTAGG) _n repeat. The presence of a telomerase coding gene has been verified successively in the A. pisum genome, revealing that aphid telomerase shares sequence identity ranging from 12% to 18% with invertebrate and vertebrate homologues, and possesses the two main domains involved in telomerase activity. Interestingly, telomerase expression has been verified in different somatic tissues suggesting that, in aphids, telomerase activity is not as restricted as in human cells. The study of telomeres in a M. persicae strain with a variable chromosome number showed that aphid telomerase can initiate the de novo synthesis of telomere sequences at internal breakpoints, resulting in the stabilization of chromosomal fragments.     

Insecticidal activity of essential oils from native medicinal plants of Central Argentina against the house fly, <Emphasis Type="Italic">Musca domestica</Emphasis> (L.)

The insecticidal activity of nine essential oils (EOs) against the house fly (Musca domestica) was evaluated by placing flies in a screw-cap glass jar holding a piece of EO-treated cotton yarn. The dose necessary to kill 50% of flies (LC₅₀) in 30 min was determined at 26?±?1°C. The EOs showed LC₅₀ values ranging from 0.5 to 46.9 mg/dm³. The EO from Minthostachys verticillata was the most potent insecticide (LC₅₀?=?0.5 mg/dm³) followed by EOs from Hedeoma multiflora (LC₅₀?=?1.3 mg/dm³) and Artemisia annua (LC₅₀?=?6.5 mg/dm³). The compositions of the nine EOs, obtained by hydrodistillation of medicinal herbs, were analyzed by gas chromatography/mass spectroscopy. These analyses showed that (4R)(+)-pulegone (69.70%), menthone (12.17%), and limonene (2.75%) were the principal components of M. verticillata EO. (4R)(+)-pulegone was also the main constituent (52.80%) of H. multiflora, while artemisia ketone (22.36%) and 1,8-cineole (16.67%) were the major constituents of A. annua EO. The terpene (4R)(+)-pulegone showed a lower toxicity (LC₅₀?=?1.7 mg/dm³) than M. verticillata or H. multiflora EOs. Dimethyl 2,2-dichlorovinyl phosphate, selected as a positive control, showed an LC₅₀ of 0.5 mg/dm³. EOs from M. verticillata and H. multiflora show promise as natural insecticides against houseflies.     

Anti-staphylococcal,anti-HIV and cytotoxicity studies of four South African medicinal plants and isolation of bioactive compounds from <Emphasis Type="Italic">Cassine transvaalensis</Emphasis> (Burtt. Davy) codd

Background

Medicinal plants represent an important opportunity to rural communities in Africa, as a source of affordable medicine and as a source of income. Increased patient awareness about safe usage is important as well as more training with regards to traditional medicine. The aim of this study was to evaluate the ethnomedicinal prowess of some indigenous South African plants commonly used in Eastern Cape Province of South Africa for the treatment of skin and respiratory tract infections, HIV and their toxicity potential.

Methods

Cassine transvaalensis, Vangueria infausta, Croton gratissimus and Vitex ferruginea were tested for antibacterial activities against Staphylococcus aureus and Staphylococcus epidermidis using Kirby-Bauer disk diffusion and minimum inhibition concentration (MIC). Cytotoxic and anti-HIV-1 activities of plants were tested using MTT Assay (3- (Dimethylthiozole-2-yl-2,5-diphenyltetrazolium bromide)) and anti- HIV-1iib assay. In search of bioactive lead compounds, Cassine transvaalensis which was found to be the most active plant extract against the two Staphylocoous bacteria was subjected to various chromatographic. Thin layer chromatography, Column chromatography and Nuclear Magnetic Resonance (NMR), (¹H-¹H, ¹³C-¹³C, in DMSO_d6, Bruker 600 MHz) were used to isolate and characterize 3-Oxo-28-hydroxylbetuli-20(29)-ene and 3,28-dihydroxylbetuli-20(29)-ene bioactive compounds from C. transvaalensis.

Results

The four plants studied exhibited bioactive properties against the test isolates. The zones of inhibition ranged between 16 mm to 31 mm for multi-drug resistant staphylococci species. MIC values varied between 0.6 and 0.02 μg/ml. C. gratissimus and C. transvaalensis exhibited the abilities to inhibit HIV-1_iib. Two bioactive compounds were isolated from C. transvaalensis.

Conclusion

Data from this study reveals the use of these plant by traditional healers in the Eastern Cape. Furthermore, C. transvaalensis and C. gratissimus were found to be more active as against HIV-1iib. While C. transvaalensis was most active against the two Staphylococcus bacteria.