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Acquiring resistance against antiviral drugs is a significant problem in antimicrobial therapy. In order to identify novel antiviral compounds, the antiviral activity of eight plants indigenous to the southern region of Hungary against herpes simplex virus-2 (HSV-2) was investigated. The plant extracts and the plant compound carnosic acid were tested for their effectiveness on both the extracellular and intracellular forms of HSV-2 on Vero and HeLa cells. HSV-2 replication was measured by a direct quantitative PCR (qPCR). Among the tested plant extracts, Salvia rosmarinus ( S. rosmarinus ) exhibited a 90.46% reduction in HSV-2 replication at the 0.47 μg/mL concentration. Carnosic acid, a major antimicrobial compound found in rosemary, also demonstrated a significant dose-dependent inhibition of both extracellular and intracellular forms of HSV-2. The 90% inhibitory concentration (IC 90 ) of carnosic acid was between 25 and 6.25 μg/mL. Proteomics and high-resolution respirometry showed that carnosic acid suppressed key ATP synthesis pathways such as glycolysis, citrate cycle, and oxidative phosphorylation. Inhibition of oxidative phosphorylation also suppressed HSV-2 replication up to 39.94-fold. These results indicate that the antiviral action of carnosic acid includes the inhibition of ATP generation by suppressing key energy production pathways. Carnosic acid holds promise as a potential novel antiviral agent against HSV-2.

Background: Earlier studies reported alterations of the kynurenine (KYN) pathway of tryptophan (TRP) metabolism in Parkinson's disease (PD). The first rate-limiting enzymes indoleamine 2,3-dioxygenase (IDO) and tryptophan dioxygenase were observed upregulated, resulting elevated KYN/TRP ratios in the serum and cerebrospinal fluid samples of patients with PD. More and more single nucleotide polymorphisms (SNPs) have been identified in a population of PD. However, little is known about the impact of genetic variations of the IDO on the pathogenesis of PD., Methods: SNP analysis of IDO1 was performed by allelic discrimination assay with fluorescently labelled TaqMan probes and a subgroup analysis was conducted according to the age of PD onset. The frame shifts variant rs34155785, intronic variant rs7820268, and promotor region variant rs9657182 SNPs of 105 PD patients without comorbidity were analyzed and compared to 129 healthy controls., Results: No significant correlation was found in three SNPs between PD patients and healthy controls. However, the subgroup analysis revealed that A alleles of rs7820268 SNP or rs9657182 SNP carriers contribute to later onset of PD than non-carriers., Conclusions: The study suggested that SNPs of IDO1 influenced the age onset of PD and genotyping of SNPs in certain alleles potentially serves as a risk biomarker of PD., Competing Interests: Given his role as Guest Editor, Masaru Tanaka had no involvement in the peer-review of this article and has no access to information regarding its peer-review. Full responsibility for the editorial process for this article was delegated to Graham Pawelec. The authors declare no conflict of interest., (© 2022 The Author(s). Published by IMR Press.)

Carbapenem-resistant Bacteroides fragilis strains usually emerge by an insertion sequence (IS) jump into the upstream region of the cfiA carbapenemase gene. However, intermediate or fully resistant cfiA -positive strains also exist. These do not have such IS element activations, but usually have heterogeneous resistance (HR) phenotypes, as detected by a disc diffusion or gradient tests. Heteroresistance is a serious antibiotic resistance problem, whose molecular mechanisms are not fully understood. We aim to characterize HR and investigate diagnostic issues in the set of cfiA -positive B. fragilis strains using phenotypic and molecular methods. Of the phenotypic methods used, the population analysis profile (PAP) and area under curve (AUC) measurements were the best prognostic markers for HR. PAP AUC, imipenem agar dilution and imipenemase production corresponded well with each other. We also identified a saturation curve parameter (quasi-PAP curves), which correlated well with these phenotypic traits, implying that HR is a stochastic process. The genes, on a previously defined ' cfiA element', act in a complex manner to produce the HR phenotype, including a lysine-acetylating toxin and a lysine-rich peptide. Furthermore, imipenem HR is triggered by imipenem. The two parameters that most correlate with the others are imipenemase production and 'GNAT' expression, which prompted us to suspect that carbapenem heteroresistance of the B. fragilis strains is stochastically regulated and is mediated by the altered imipenemase production.

Urinary tract infections (UTIs) are common bacterial infections caused mainly by enteric bacteria. Numerous virulence factors assist bacteria in the colonization of the bladder. Bacterial efflux pumps also contribute to bacterial communication and to biofilm formation. In this study, the phenotypic and genetic antibiotic resistance of clinical UTI pathogens such as Escherichia coli , Klebsiella pneumoniae , and Proteus mirabilis were determined by disk diffusion method and polymerase chain reaction (PCR). Following this, different classes of antibiotics were evaluated for their antibacterial activity at pH 5, 6, 7 and 8 by a microdilution method. Gentamicin (GEN) was the most potent antibacterial agent against E. coli strains. The effect of GEN on the relative expression of marR and sdiA genes was evaluated by quantitative PCR. The slightly acidic pH (pH 6) and GEN treatment induced the upregulation of marR antibiotic resistance and sdiA QS activator genes in both E. coli strains. Consequently, bacteria had become more susceptible to GEN. It can be concluded that antibiotic activity is pH dependent and so the artificial manipulation of urinary pH can contribute to a more effective therapy of multidrug resistant bacterial infections.

Eleven metronidazole resistant Bacteroides and one newly classified Phocaeicola dorei strain from Kuwait were investigated for their resistance mechanisms and the emergence of their resistant plasmids. All but one strain harbored nimE genes on differently sized plasmids. Of the 11 nimE genes, 9 were preceded by full copies of the prototype ISBf6 insertion sequence element, one carried a truncated ISBf6 and one was activated by an additional copy of IS612B. Nucleotide sequencing results showed that the nimE ISBf6 distances were constant and all five different plasmids shared a common region, suggesting that (i) the nimE-ISBf6 configuration was inserted into an undisclosed common genetic element, (ii) over time, this common element was mutated by insertions and deletions, spreading the resultant plasmids. Of the 10 B. fragilis strains in this collection, 6 were also cfiA-positive, one with full imipenem resistance, indicating a tendency for multidrug resistance (MDR) among such isolates. The significant number of metronidazole resistant Bacteroides spp. and P. dorei strains with the MDR phenotype warns of difficulties in treatment and suggests promoting adherence to antibiotic stewardship recommendations in Kuwait., Competing Interests: Declaration of competing interest We declare no conflict of interests., (Copyright © 2021 Elsevier Ltd. All rights reserved.)

Purpose of Review: SARS-CoV-2, the recently emerged coronavirus (CoV) that is responsible for the current global pandemic Covid-19, first appeared in late 2019 in Wuhan, China. Here, we summarise details of the SARS-CoV-2 genome to assist understanding of the emergence, evolution and diagnosis of this deadly new virus., Recent Findings: Based on high similarities in the genome sequences, the virus is thought to have arisen from SARS-like CoVs in bats but the lack of an intermediate species containing a CoV with even greater similarity has so far eluded discovery. The critical determinant of the SARS-CoV-2 genome is the spike (S) gene encoding the viral structural protein that interacts with the host cell entry receptor ACE2. The S protein is sufficiently adapted to bind human ACE2 much more readily than SARS-CoV, the most closely related human CoV., Summary: Although the SARS-CoV-2 genome is undergoing subtle evolution in humans through mutation that may enhance transmission, there is limited evidence for attenuation that might weaken the virus. It is also still unclear as to the events that led to the virus' emergence from bats. Importantly, current diagnosis requires specific recognition and amplification of the SARS-CoV-2 RNA genome by qPCR, despite these ongoing viral genome changes. Alternative diagnostic procedures relying on immunoassay are becoming more prevalent., Competing Interests: Conflict of InterestThe authors declare that they have no conflict of interest., (© The Author(s), under exclusive licence to Springer Science+Business Media, LLC part of Springer Nature 2021.)

Objective: Herpes simplex virus-2 (HSV-2) infections are almost exclusively sexually transmitted. The presence of vaginal gels during sexual activity may have a significant positive or negative impact on viral transmission. Therefore we investigated three off-the-shelf vaginal lubricants and one pH restoring gel to evaluate their impact on HSV-2 replication., Results: HeLa cells were infected with untreated virions and virions incubated with the particular gels. The accumulation of viral genomes was monitored by quantitative PCR (qPCR) method at 24 h post infection. Two of the tested gels had no significant effect on HSV-2 replication at the maximum applied concentration, while two had a strong inhibitory effect (~ 98% reduction of replication). The replication inhibitory effect was observed at various multiplicity of infection (MOI 0.4-6.4) and the two inhibitory gels were also capable of inhibiting the HSV-2 induced cytopathic effect on HeLa cells. The surface tension decreasing activity-an indication of detergent activity-was strongly correlated with the anti-HSV-2 activity of the gels (R 2 : 0.88). Our results indicate that off-the-shelf vaginal gels have a markedly different anti-HSV-2 activity that may influence HSV-2 transmission.

Clinical studies have shown that schizophrenia is accompanied by hypoalgesia. Accordingly, we have previously reported that a chronic schizophrenia-related rat substrain (Wisket) showed decreased acute heat pain sensitivity. The aim of the present study was to determine the mechanical pain sensitivity and the effects of opioid ligands in a chronic osteoarthritic pain model generated using Wisket rats. Our previous molecular biological studies indicated that the impairment in opioid and cannabinoid receptor functions observed in these animals did not explain their altered pain sensitivity. Therefore, we aimed to investigate another endogenous antinociceptive system, i.e., the oxytocinergic system (which is also implicated in schizophrenia) via the determination the brain-region specific oxytocin receptor mRNA expression in Wisket rats. Osteoarthritis was induced in male adult control Wistar rats without any interventions and in Wisket rats after juvenile social isolation and ketamine treatment. The degree of allodynia and the effects of systemic morphine or intrathecal endomorphin-1 administration were determined. Furthermore, the expression of the oxytocin receptor mRNA was assessed in different brain structures (prefrontal cortex, striatum, diencephalon, brainstem, and olfactory bulb). A lower degree of allodynia was observed in the Wisket group compared with control animals 1 and 2 weeks after the induction of osteoarthritis, which was accompanied by a comparable degree of edema. Systemically or intrathecally applied opioids caused similar time-response curves in both groups, with apparently shorter effects in Wisket animals. The expression of the oxytocin receptor mRNA was lower in most of the brain regions (with the exception of the diencephalon) investigated in Wisket rats vs. the control animals. In summary, both acute and chronic hypoalgesia (as nonspecific symptoms in patients with schizophrenia) can be simulated in Wisket animals as endophenotypes despite the impairment of the endogenous antinociceptive systems evaluated. Thus, this model might be an appropriate tool for further investigation of the molecular basis of altered pain perception in schizophrenia., (Copyright © 2019 Elsevier B.V. All rights reserved.)

Objective: We aim to directly detect Mycoplasma DNA in a U937 suspension cell culture without using DNA purification. In order to make Mycoplasma contamination monitoring easier, we optimized a commercially available quantitative PCR (qPCR)-based detection kit. We compared the sensitivity of direct qPCR against qPCR with a purified DNA template., Results: Our findings indicate that qPCR worked optimally with a 6 μl sample volume and a 52 °C annealing-extension temperature. We were able to decrease the annealing-extension step time from 60 to 20 s without any major decrease in reaction sensitivity. The total cycle time of optimized direct qPCR was 65 min. The optimized qPCR protocol was used to detect Mycoplasma DNA before and after DNA purification. Our findings indicate that direct qPCR had a higher sensitivity than regular qPCR. Ct levels produced by direct qPCR with 6 μl templates were almost identical to Ct levels produced by regular qPCR with DNA purified from a 60 μl cell culture sample (23.42 vs 23.49 average Ct levels, respectively). The optimized direct qPCR protocol was successfully applied to monitor the elimination of Mycoplasma contamination from U937 cell cultures.

Chlamydia trachomatis infections are the most prevalent sexually transmitted infections with potentially debilitating sequelae, such as infertility. Mouse models are generally used for vaccine development, to study the immune response and histopathology associated with Chlamydia infection. An important question regarding murine models is the in vivo identification of murine host genes responsible for the elimination of the murine and human Chlamydia strains. RNA sequencing of the Chlamydia muridarum infected BALB/c lung transcriptome revealed that several genes with direct antichlamydial functions were induced at the tissue level, including the already described and novel members of the murine interferon-inducible GTPase family, the CXCL chemokines CXCL9, CXCL11 , immunoresponsive gene 1, nitric oxide synthase-2 ( iNOS ), and lipocalin-2. Indoleamine 2,3-dioxygenase 1-2 ( IDO1-2 ) previously described potent antichlamydial host enzymes were also highly expressed in the infected murine lungs. This finding was novel, since IDO was considered as a unique human antichlamydial defense gene. Besides a lower level of epithelial cell positivity, immunohistochemistry showed that IDO1-2 proteins were expressed prominently in macrophages. Detection of the tryptophan degradation product kynurenine and the impact of IDO inhibition on Chlamydia muridarum growth proved that the IDO1-2 proteins were functionally active. IDO1-2 activity also increased in Chlamydia muridarum infected C57BL/6 lung tissues, indicating that this phenomenon is not mouse strain specific. Our study shows that the murine antichlamydial response includes a variety of highly up-regulated defense genes in vivo . Among these genes the antichlamydial effectors IDO1-2 were identified. The potential impact of murine IDO1-2 expression on Chlamydia propagation needs further investigation.

The DEFB4 gene copy numbers were investigated in 206 AD patients and in 250 controls. The levels of the human defensin β-2 (hBD2) and α-defensins (HNP 1-3) in the sera and in the cerebrospinal fluid (CSF) of the patients and the controls were determined. Higher copy numbers of the DEFB4 gene was observed in AD patients as compared with the controls. The levels of hBD-2 and HNP 1-3 were significantly elevated in the sera and in the CSF of the AD patients These data suggest that both defensin β-2 and α-defensins have potential role in the development of AD., (Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.)

Chlamydiae are obligate intracellular bacteria developing in an intracytoplasmic niche, the inclusion. Chlamydia growth measurement by inclusion counting is a key task in the development of novel antichlamydial antibiotics and in vaccine studies. Most of the current counting methods rely on the immunofluorescent staining of the inclusions and either manual or automatic microscopy detection and enumeration. The manual method is highly labor intensive, while the automatic methods are either medium-throughput or require automatic microscopy. The sensitive and specific PCR technology could be an effective method for growth related chlamydial DNA detection; however the currently described PCR approaches have a major limitation, the requirement of purification of DNA or RNA from the infected cells. This limitation makes this approach unfeasible for high-throughput screenings. To overcome this, we developed a quantitative PCR (qPCR) method for the detection of Chlamydia trachomatis DNA directly from the infected HeLa cells. With our method we were able to detect the bacterial growth in a 4 log scale (multiplicity of infection (MOI): 64 to 0.0039), with high correlation between the biological and technical replicates. As a further proof of the method, we applied the direct qPCR for antibiotic minimum inhibitory concentration (MIC) measurements. The measured MICs of moxifloxacin, tetracycline, clarithromycin and compound PCC00213 were 0.031 μg/ml, 0.031 μg/ml, 0.0039 μg/ml and 6.2 μg/ml respectively, identical or close to the already published MIC values. Our direct qPCR method for chlamydial growth and antibiotic MIC determination is less time-consuming, more objective and more sensitive than the currently applied manual or automatic fluorescent microscopy- based methods., (Copyright © 2015 Elsevier B.V. All rights reserved.)

Aims: Recent studies have started to elucidate the contribution of microbiome to the pathogenesis of multiple sclerosis (MS). It is also supposed, that neuropathological alterations might be associated with abnormal expression and regulatory function of antimicrobial peptides (AMPs), including defensins. It is in our interest to investigate the relevance of the single nucleotide polymorphisms (SNPs) of the DEFB1 gene and the copy number polymorphism of the DEFB4 genes in MS., Methods: DEFBI polymorphisms: c.-20G > A (rsl 1362), DEFB1 c.-44C > G (rsI 800972), DEFB1 c.-52G>A (rsl 799946), and the DEFB4 gene copy number were investigated in 250 MS patients The control patients comprised 232 age- and gender-matched healthy blood donors. The occurrence of the human β-defensin 2 peptide (hBD2) in the plasma of controls and patients-was determined by ELISA., Results: The DEFB1 c.-44C>G polymorphism the GG protective genotype was much less frequent among patients than among the controls. A higher frequency of a lower (<4) copy number of the DEFB4 gene was observed in the patients with MS as compared with the controls (43% vs. 28%, respectively). The median levels of the circulating hBD2 in the patients were 150.6 +/- 12.71 pg/ml vs. 262.1 +/- 23.82 pg/mI in the control group (p<0.0001). Our results suggest that β-defensins play role in the development of MS.

The roles of chemokine receptor V (CCR5) and its polymorphism, rs333 in multiple sclerosis (MS) are controversial. We investigated the receptor and its deletion in a large MS (428) and a numerous control (831) population in Csongrád County (Hungary) and North-Bácska (Serbia). Taqman probes firstly were used for the allele discrimination. There was no significant difference in genotype (OR=1.092, 95% CI=0.807-1.478, p=0.568 for wt/wt (wt=wild type allele) vs wt/Δ32, Δ32/Δ32 (Δ32=Δ32 base pair deletion allele)) or allele frequency (OR=0.914, 95% CI=0.692-1.207, p=0.525). Neither the deletion nor the wt allele affected the Expanded Disability Status Scale score or the age at onset. Our results indicate no association between the CCR5 Δ32 allele and MS., (Copyright © 2014 American Society for Histocompatibility and Immunogenetics. Published by Elsevier Inc. All rights reserved.)

Background. There is substantial evidence that the kynurenine pathway (KP) plays a role in the normal physiology of the brain and is involved in the pathology of neurodegenerative disorders such as Huntington's disease and Parkinson's disease (PD). Objective. We set out to investigate the potential roles in PD of single nucleotide polymorphisms (SNPs) from one of the key enzymes of the KP, kynurenine 3-monooxygenase (KMO). Methods. 105 unrelated, clinically definitive PD patients and 131 healthy controls were enrolled to investigate the possible effects of the different alleles of KMO. Fluorescently labeled TaqMan probes were used for allele discrimination. Results. None of the four investigated SNPs proved to be associated with PD or influenced the age at onset of the disease. Conclusions. The genetic link between the KP and PD is still missing. The investigated SNPs presumably do not appear to influence the function of KMO and probably do not contain binding sites for regulatory proteins of relevance in PD. This is the first study to assess the genetic background behind the biochemical alterations of the kynurenine pathway in PD, directing the attention to this previously unexamined field.

Aim: To investigate the genetic background of human defensin expression in type 1 and 2 diabetes., Methods: Associations between DEFA1/DEFA3 gene copy number polymorphism and diabetes as well as between the promoter polymorphisms of DEFB1 and diabetes were studied. The copy number variation of the DEFA1/DEFA3 genes was determined in 257 diabetic patients (117 patients with type 1 and 140 with type 2 diabetes). The control group consisted of 221 age- and gender-matched healthy blood donors. The cumulative copy numbers of the DEFA1/DEFA3 genes were detected by using quantitative PCR analysis. To evaluate the HNP 1-3 (human neutrophil peptide 1-3 or α-defensin) levels in the circulation, plasma HNP 1-3 concentrations were measured by ELISA. The expression of DEFA1/A3 in peripheral leukocytes of the diabetic patients was measured by quantitative RT PCR analysis. Three SNPs of the human DEFB1 (human defensin β-1) gene: DEFB1 G-20A (rs11362), DEFB1 C-44G (rs1800972) and DEFB1 G-52A (rs1799946) were genotyped by Custom TaqMan(®) Real Time PCR assay., Results: Significant differences were observed in HNP1-3 levels between the healthy subjects and both groups of diabetic patients. The mean ± SE was 28.78 ± 4.2 ng/mL in type 1 diabetes, and 29.82 ± 5.36 ng/mL in type 2 diabetes, vs 11.94 ± 2.96 ng/mL in controls; P < 0.01 respectively. There was no significant difference between patients with type 1 and type 2 diabetes in the high plasma concentrations of HNP1-3. The highest concentrations of α-defensin were found in diabetic patients with nephropathy (49.4 ± 4.8 ng/mL), neuropathy (38.7 ± 4.8 ng/mL) or cardiovascular complications (45.6 ± 1.45 ng/L). There was no significant difference in the cumulative copy numbers of DEFA1/DEFA3 genes between controls and patients, or between patients with the two types of diabetes. Comparisons of HNP 1-3 plasma level and DEFA1/A3 copy number of the same patient did not reveal significant relationship between defensin-α levels and the gene copy numbers (r (2) = 0.01). Similarly, no positive correlation was observed between the copy numbers and the mRNA expression levels of DEFA1/A3. Regarding the C-44G polymorphism of DEFB1, the GG "protective" genotype was much less frequent (1%-2%) among both groups of patients than among controls (9%)., Conclusion: Elevated HNP1-3 levels in diabetes are independent of DEFA1/DEFA3 copy numbers, but GG genotype of C-44G SNP in DEFB1 gene may result in decreased defensin β-1 production.

Background: Polymerase chain reaction (PCR)-based techniques are widely used to identify fungal and bacterial infections. There have been numerous reports of different, new, real-time PCR-based pathogen identification methods although the clinical practicability of such techniques is not yet fully clarified.The present study focuses on a novel, multiplex, real-time PCR-based pathogen identification system developed for rapid differentiation of the commonly occurring bacterial and fungal causative pathogens of bloodstream infections., Results: A multiplex, real-time PCR approach is introduced for the detection and differentiation of fungi, Gram-positive (G+) and Gram-negative (G-) bacteria. The Gram classification is performed with the specific fluorescence resonance energy transfer (FRET) probes recommended for LightCycler capillary real-time PCR. The novelty of our system is the use of a non-specific SYBR Green dye instead of labelled anchor probes or primers, to excite the acceptor dyes on the FRET probes. In conjunction with this, the use of an intercalating dye allows the detection of fungal amplicons.With the novel pathogen detection system, fungi, G + and G- bacteria in the same reaction tube can be differentiated within an hour after the DNA preparation via the melting temperatures of the amplicons and probes in the same tube., Conclusions: This modified FRET technique is specific and more rapid than the gold-standard culture-based methods. The fact that fungi, G + and G- bacteria were successfully identified in the same tube within an hour after the DNA preparation permits rapid and early evidence-based management of bloodstream infections in clinical practice.

The pathogenesis of multiple sclerosis (MS), a devastating neuroinflammatory disorder of the central nervous system, has been presumed to involve the possible importance of the receptor for advanced glycation end products (RAGE). The aim of this study was to investigate the relevance of the genetic polymorphisms of RAGE in MS patients. A total of 168 patients with MS were enrolled; 136 healthy blood donors served as controls. The -374 T/ A, -479 T/C, and the G82S polymorphisms of RAGE were determined by restriction fragment length polymorphism (RFLP). There was a significant difference in RAGE -374 T/A genotype distribution between the controls and the MS patients. The AA homozygote variants were detected in 8% of the patients with MS, as compared with 19% of healthy controls (OR=2.75; 95% CI=1.319-5.733, p = 0.007). No differences were observed between the MS patients and the controls, concerning the frequencies of the -479 T/C and G82S genotypes of the RAGE. Our results revealed an association between the -374 T/A polymorphism of the RAGE promoter and MS. The genetic variant -374 AA (which has previously been shown to exert significant effects on transcriptional activity) can be considered a preventive factor as regards the occurrence of MS. Our findings support the view that RAGE plays a role in the development of MS.

In this study, the gene hmgR encoding the 3-hydroxy-3-methylglutaryl-coenzyme A reductase (HMG-CoA reductase) was cloned and characterized in the zygomycete fungus Rhizomucor miehei. The hmgR gene comprises a total of 3,585 bp including the coding sequence of a 1,058 amino acids length putative protein and five introns (137, 83, 59, 60 and 69 bp in length) dispersed in the whole coding region. Southern hybridization analysis revealed that the gene is present only in one copy in the R. miehei genome. The isolated Rhizomucor gene was expressed in the related fungus, Mucor circinelloides. Transformants harbouring the Rhizomucor hmgR gene in an autoreplicative plasmid proved to be more tolerant to statins (e.g. lovastatin, simvastatin, and fluvastatin), the competitive inhibitors of the HMG-CoA reductase, than the original M. circinelloides strain. At the same time, heterologous expression of the Rhizomucor hmgR did not affect the carotenoid production of M. circinelloides.

Objective: It has been suggested that deficient defensin expression is associated with the chronic inflammation of Crohn's disease. The regional localization of Crohn's disease, ileal or colonic disease can be linked to different defensin profiles. As constitutive beta-defensin 1 has a colonic expression, we considered it of interest to investigate single nucleotide polymorphisms (SNPs) of the beta-defensin 1 gene (DEFB1) in Crohn's disease., Material and Methods: Three SNPs of the DEFB1 gene, DEFB1 G-20A (rs11362), DEFB1 C-44G (rs1800972) and DEFB1 G-52A (rs1799946), were genotyped either by Custom TaqMan SNP genotyping assays or by restriction fragment length polymorphisms (RFLP) in 190 patients with Crohn's disease and 95 Hungarian controls., Results: It was found that the G-20A and C-44G SNPs had a strong association with the colonic and ileocolonic localizations of the disease, respectively, but no association was detected for the ileal localization. A significantly higher frequency of the GA genotype of G-20A was observed among patients with colonic localization (60%) as compared with healthy controls (39%), with an odds ratio (OR) of 2.39. The GG genotype of C-44G SNP, which is regarded as a protective genotype, was much less frequent (4%) among patients than among controls (12%), OR 3.367., Conclusions: These results indicate that genetic variations in the DEFB1 gene encoding constitutive human beta-defensin 1 may be associated with the risk for Crohn's disease and may determine disease phenotype, e.g. colonic localization.

The authors present the cases of two infants with congenital cytomegalovirus infection, who to the best of the authors' knowledge were the first ones to receive ganciclovir treatment in Hungary. Both infants were treated for symptomatic congenital cytomegalovirus infection affecting the central nervous system. Ganciclovir was given intravenously in two phases, for a total of 4 and 6 weeks, in a daily dose of 5-10 mg/kg. Diagnosis of infection and follow-up of treatment efficacy was based on the quantitative assessment by PCR assay of viral nucleic acid in blood and urine samples. Treatment resulted in substantial reduction of viral copy numbers in both infants' blood and urine samples. Improvement in the biochemical markers of disease activity was accompanied with spectacular improvement of clinical symptoms in the infant with severe liver involvement. Following the treatment viral loads increased in both infant but clinical symptoms did not reoccur. In one patient a considerable improvement of hearing loss was observed. The authors' first results indicate that ganciclovir treatment of neonatal cytomegalovirus infection represents a promising approach in preventing the progression of the disease and in ameliorating the consequences.

Introduction: Methylenetetrahydrofolate reductase (MTHFR) C677T and A1298C mutations, being considered unfavourable genetic factors by causing elevated serum homocysteine levels, may be risk factors for cardiovascular disorders, including ischaemic stroke. In this study, the role of these two mutations in ischaemic stroke was examined:, Patients and Methods: Genetic and clinical data were analysed of 122 ischaemic stroke patients and 102 control subjects with no lesions by neuroimaging., Results: Neither of the two MTHFR mutations alone was found to be a significant genetic risk factor for ischaemic stroke. However, at least one MTHFR 677T allele combined with at least one MTHFR 1298C allele significantly increased the risk of ischaemic stroke (adjusted odds ratio: 3.39; p < 0.001)., Conclusion: The synergistic effect between the two MTHFR mutations may represent a new genetic stoke risk factor.

Recent observations revealed a novel role of angiotensin-converting enzyme 2 and the angiotensin II type-1 receptor (AT1R) in lung injury, thereby extending knowledge about the functions of the angiotensin system. Angiotensin II, whose target is the AT1R, is a potent vasoconstrictor. Accordingly, an imbalance leading to enhanced activity of the angiotensin II-AT1R axis is postulated to contribute to both circulatory disturbances and lung injury. In this context, a functional single-nucleotide polymorphism, AT1R A1166C, which leads to enhanced responsiveness of the AT1R, has been postulated as a candidate susceptibility factor for ischemic stroke. The aim of our study was to investigate its occurrence in ischemic stroke and to analyze its possible synergistic associations with clinical risk factors. Genetic and clinical data on 308 consecutive patients with acutely developing ischemic stroke were analyzed. A total of 272 stroke and neuroimaging alteration-free subjects served as a control group. Univariate and logistic regression statistical approaches were used. Alone, the AT1R 1166C allele did not pose a risk of stroke. In hypertensive smokers, however, it was associated with an increased risk of ischemic stroke (OR 22.3, 95% CI 5.8-110.2, p<0.001). Further subgroup analysis revealed the same association for both small-vessel (OR 24.3, 95% CI 6.1-121.1, p<0.001) and large-vessel (OR 21.3, 95% CI 4.6-81.1, p<0.001) infarction. On a pathophysiological basis, our results suggest the possibility that the AT1R A1166C polymorphism might give rise to ischemic stroke indirectly via an unfavorable effect on the cardiorespiratory function.

The possible pathogenic role of triglycerides (TG) in the development of ischemic stroke is still under extensive investigation. Recently, apolipoprotein (apo)A5 gene promoter region T-1131C polymorphism has been shown to associate with elevated serum TG levels. In the current work, a total of 302 subjects were classified as being large vessel-associated, small vessel-associated, or belonging to a mixed group of ischemic stroke-affected patients. The level of TG was increased in all groups (p < 0.01). The apoA5-1131C allele frequency was approximately twofold in all groups of stroke patients compared with the controls (5 vs 10-12%; p < 0.05); and the apoA5-1131C allele itself was also found to associate with increased TG levels in all groups. In a multivariate logistic regression analysis model adjusted for differences in age, gender, serum cholesterol, hypertension, presence of diabetes mellitus, smoking and drinking habits, and ischemic heart disease, a significantly increased risk of developing stroke disease was found in patients carrying the apoA5-1131C allele (p < 0.05; odds ratio OR = 2.1 [1.3-4.7]); this association was also proven for all subtypes of the stroke. The results presented here suggest that the apoA5-1131C allele is an independent risk factor for the development of stroke. Being that apoA5 gene is under the control of the peroxisome proliferator-activated receptor alpha, theoretically, the current observations also can have long-term therapeutic consequences.

A direct role of lymphotoxin-alpha (LTA) in promoting atherosclerotic plaque growth has been demonstrated recently. The different protein transcripts of the naturally occurring genetic variants of the LTA gene have been demonstrated to exhibit affected functions, and an allelic difference in binding to transcription factor(s) has also been suggested. The homozygous variant of LTA characterized by the intron 1 252A-->G (252G) transition, which naturally coexists with an exon 3 804C-->A (804A) single-nucleotide polymorphism (SNP), has been reported as a susceptibility gene for myocardial infarction. Because the atherosclerotic process is also an integral component in the pathogenesis of certain types of vascular stroke, we investigated the possible significance of the above SNPs in 353 ischemic stroke patients and 180 healthy controls. The homozygous LTA allele with the 252G and 804C SNPs occurred more frequently in stroke patients (13.9%) than in controls (7.20%, p<0.025). Specific subclassification of the patients revealed an accumulation of these SNPs in large-vessel, pathology-associated cerebral infarction (18.2%); multivariate logistic regression analysis of the data confirmed this association, with an odds ratio of 2.1 (95% confidence interval, 1.3-6.2; p<0.005). Elimination of all subjects with a history or evidence of ischemic heart disease, including myocardial infarction, did not affect this association. These data show that besides the role in the development of myocardial infarction, the homozygous carriage of the LTA allele with 252G and 804A SNPs is a novel susceptibility factor for largevesselassociated ischemic stroke.

Introduction: Platelet glycoprotein IIb/IIIa is a membrane receptor with a central function in the platelet adhesion and ultimately in the thrombus formation. Two major variants of the gene encoding the IIIa subunit, called PLA1 (A1) and PLA2 (A2), have been identified in the general population. There are indications that the A2 allele can also be associated with acute thrombosis or stroke. The purpose of this study was to study the distribution of the A2 allele in different vascular subtypes of stroke disease., Materials and Methods: A total of 638 consecutive patients were analyzed and classified as having large vessel pathology (n=168) or a small vessel infarct (n=210). Localization of the vascular occlusions was deducted from analysis of the magnetic resonance imaging (MRI) scan results in stroke patients. The remainder patients were listed into a mixed vascular pathology group (n=167). Patients with other or poorly characterized stroke etiology were excluded from the study (n=93)., Results: In the small vessel and mixed vascular pathology groups, the PLA2 allele frequency was similar to that in the controls. By contrast, PLA2 allele frequency was approximately two-fold higher in patients with large vessel pathology (23.3%) than in the stroke-free control subjects (11.7%, p<0.0005). Multivariate logistic regression analysis of data confirmed this association with an odds ratio (OR) of 2.9 (95% confidence interval [CI]: 1.6-4.9, p<0.0005)., Conclusions: These data suggest that the PLA2 allele is more frequent in brain infarcts associated with large-vessel occlusion.

Objective: Ischemic stroke is a frequent heterogeneous multifactorial disease that is affected by several genetic mutations and environmental factors. We hypothesised the clinical importance of the co-occurrence of common, unfavorable genetic mutations in the development of different stroke subtypes., Method and Material: The Factor V Leiden G1691A (Leiden V), the prothrombin G20210A and the methylenetetrahydrofolate reductase C677T (MTHFR C677T) mutations and the angiotensin-converting enzyme I/D (ACE I/D) and apolipoprotein E (APOE) genotypes were examined by the PCR technique in 689 ischemic stroke patients and 652 stroke-free controls. Logistic regression models were used to estimate the relative risks of different stroke subtypes for different genotype combination patterns., Results: The ACE D/D genotype alone or in combination with the MTHFR 677T or the APOE 4 allele or with both was highly specific for small-vessel infarction. The Leiden V mutation alone or in different combination patterns with the ACE D, APOE 4 and MTHFR 677T alleles was specifically predisposed to large-vessel infarction. The APOE 4 allele alone was calculated to be a general, minor genetic risk factor for ischemic stroke. The MTHFR 677T allele alone was not a risk factor for any stroke subtype. In the different specific predisposition gene combinations, however, both the APOE 4 and MTHFR 677T alleles could increase the relative risk of the given stroke subgroup., Conclusions: Common mutations which alone are minor or non-significant risk factors for ischemic stroke can yield, in specific combination patterns, a highly significant, moderate genetic risk of specific stroke subtypes.