Development and in vitro antiviral activity of ivermectin liposomes as a potential drug carrier system.

This study aimed to assess and compare diverse formulations of ivermectin-loaded liposomes, employing lipid film hydration and ethanol injection methods. Three lipids (DOPC, SPC, and DSPC) were used in predetermined molar ratios. A total of 18 formulations were created, and a factorial design determined the optimal formulation based on particle size, polydispersity index (PDI), zeta potential, and encapsulation efficiency. The average mean particle size, PDI and zeta potential of the selected formulations (F1, F2, F7, F9, and F11) was, respectively, 196.40 ± 44.60 nm, 0.39 ± 0.09, and -40.24 ± 9.17. The encapsulation efficiency exceeded 80%, with a mean loading capacity of 4.00 ± 1.70%. In vitro studies included transmission electron microscopy, Fourier transform infrared spectroscopy, drug release, and antiviral activity assessments against SARS-CoV-2. The liposomal formulations demonstrated superior antiviral activity compared to free ivermectin, as indicated by lower IC50 values. The results of this study emphasize the effectiveness of ivermectin-loaded liposomes in inhibiting viral activity, highlighting their potential as promising candidates for antiviral therapy. The findings suggest that the strategic use of liposomes as drug carriers can significantly modulate and improve the antiviral properties of ivermectin, offering a novel approach to harnessing its full therapeutic potential. Collectively, these results provide a robust foundation for further exploration of ivermectin as a viral protection tool and optimization of its delivery mechanisms.

[Investigation of Ganciclovir Resistance in Cytomegalovirus Isolates by Phenotypic and Genotypic Methods]. / Sitomegalovirüs Izolatlarinda Gansiklovir Direncinin Fenotipik ve Genotipik Yöntemlerle Arastirilmasi.

Ganciclovir-resistant cytomegalovirus (CMV) strains are reported following long-term antiviral agent use, especially for immune-suppressive patients. In this study, it was aimed to investigate the mutations in the UL97 gene of CMV, which causes ganciclovir (GCV) resistance by genotypic and phenotypic methods in patients who developed CMV infection following hematopoietic cell (HCT) or solid organ transplantation (SOT). Thirty patients who had HCT or SOT in Mediterranean University Hospital and developed CMV infection during routine follow-up with a viral load of CMV over 1000 copies/mL were included in the study. CMV DNA was analyzed by an automated system (Cobas Ampliprep/COBAS TaqMan CMV Test, Roche Diagnostics, Germany) quantitatively. DNA sequence analysis of the regions including codons 420-664 in the UL97 gene region was done by the Sanger sequencing method to detect mutations causing antiviral resistance and compared with defined mutations. In order to investigate antiviral resistance by phenotypic methods, heparinized blood samples of the patients were collected, 'buffy coat (leukocyte layer)' was inoculated into MRC-5 cells by centrifugation method and CMV growth in these cells was controlled with monoclonal antibodies when growth was detected, virus titer was determined and plaque reduction test was applied as recommended. It was determined that 22 of the 30 patients were HCT recipients and eight were SOT (five kidney, three liver) recipients. When the CMV serology pattern of the patients was evaluated before transplantation, 29 (96.7%) patients were found to be seropositive and one (3.3%) patient was found to be seronegative. Totally, nine CMV UL97 mutations were detected in seven (23.3%) pediatric patients who had HCT, including six seropositive and one seronegative case. In addition, one mutation (D605E) not known to cause GCV resistance was detected in a seronegative recipient and three previously unidentified mutations were detected (1474T, F499S, V559A) in a seronegative recipient. Five of the mutations defined were UL97 mutations with a defined clinical resistance against GCV in each of the five recipients (C603W, C592G, H520Q, M460V, A594T). In the plaque reduction test using 3 µM, 12 µM, 48 µM and 96 µM concentrations of GCV in CMV strains, the IC50 value was determined to be ≥ 8 µM for the five CMV strains, and the phenotypic presence of GCV resistance was shown. Clinical resistance associated with CMV UL97 mutation was detected in five (22.7%) of 22 patients who had HCT. GCV resistance was also demonstrated in these patients by phenotypic methods. No UL97 mutation was detected in the patients who had SOT.

Efficacy and safety of an inactivated whole-virion SARS-CoV-2 vaccine (CoronaVac): interim results of a double-blind, randomised, placebo-controlled, phase 3 trial in Turkey.

BACKGROUND: CoronaVac, an inactivated whole-virion SARS-CoV-2 vaccine, has been shown to be well tolerated with a good safety profile in individuals aged 18 years and older in phase 1/2 trials, and provided a good humoral response against SARS-CoV-2. We present the interim efficacy and safety results of a phase 3 clinical trial of CoronaVac in Turkey. METHODS: This was a double-blind, randomised, placebo-controlled phase 3 trial. Volunteers aged 18-59 years with no history of COVID-19 and with negative PCR and antibody test results for SARS-CoV-2 were enrolled at 24 centres in Turkey. Exclusion criteria included (but were not limited to) immunosuppressive therapy (including steroids) within the past 6 months, bleeding disorders, asplenia, and receipt of any blood products or immunoglobulins within the past 3 months. The K1 cohort consisted of health-care workers (randomised in a 1:1 ratio), and individuals other than health-care workers were also recruited into the K2 cohort (randomised in a 2:1 ratio) using an interactive web response system. The study vaccine was 3 µg inactivated SARS-CoV-2 virion adsorbed to aluminium hydroxide in a 0·5 mL aqueous suspension. Participants received either vaccine or placebo (consisting of all vaccine components except inactivated virus) intramuscularly on days 0 and 14. The primary efficacy outcome was the prevention of PCR-confirmed symptomatic COVID-19 at least 14 days after the second dose in the per protocol population. Safety analyses were done in the intention-to-treat population. This study is registered with ClinicalTrials.gov (NCT04582344) and is active but no longer recruiting. FINDINGS: Among 11 303 volunteers screened between Sept 14, 2020, and Jan 5, 2021, 10 218 were randomly allocated. After exclusion of four participants from the vaccine group because of protocol deviations, the intention-to-treat group consisted of 10 214 participants (6646 [65·1%] in the vaccine group and 3568 [34·9%] in the placebo group) and the per protocol group consisted of 10 029 participants (6559 [65·4%] and 3470 [34·6%]) who received two doses of vaccine or placebo. During a median follow-up period of 43 days (IQR 36-48), nine cases of PCR-confirmed symptomatic COVID-19 were reported in the vaccine group (31·7 cases [14·6-59·3] per 1000 person-years) and 32 cases were reported in the placebo group (192·3 cases [135·7-261·1] per 1000 person-years) 14 days or more after the second dose, yielding a vaccine efficacy of 83·5% (95% CI 65·4-92·1; p<0·0001). The frequencies of any adverse events were 1259 (18·9%) in the vaccine group and 603 (16·9%) in the placebo group (p=0·0108) with no fatalities or grade 4 adverse events. The most common systemic adverse event was fatigue (546 [8·2%] participants in the vaccine group and 248 [7·0%] the placebo group, p=0·0228). Injection-site pain was the most frequent local adverse event (157 [2·4%] in the vaccine group and 40 [1·1%] in the placebo group, p<0·0001). INTERPRETATION: CoronaVac has high efficacy against PCR-confirmed symptomatic COVID-19 with a good safety and tolerability profile. FUNDING: Turkish Health Institutes Association.

The changing dynamics of neutralizing antibody response within 10 months of SARS-CoV-2 infections.

There are limited data on how long neutralizing antibody (NAb) response elicited via primary SARS-CoV-2 infection will last. Eighty-four serum samples were obtained from a prospective cohort of 42 laboratory-confirmed COVID-19 inpatients at the time of discharge from the hospital and in the late convalescent phase. A virus neutralization assay was performed to determine the presence and titers of NAbs with authentic SARS-CoV-2. Long-term dynamics of NAbs and factors that may have an impact on humoral immunity were investigated. Mild and moderate/severe patients were compared. The mean sampling time was 11.12 ± 5.02 days (4-28) for the discharge test and 268.12 ± 11.65 days (247-296) for the follow-up test. NAb response was present in 83.3% of the patients about 10 months after infection. The detectable long-term NAb rate was significantly higher in mild patients when compared to moderate/severe patients (95.7% vs. 68.4%, p = 0.025). In the follow-up, NAb-positive and -negative patients were compared to determine the predictors of the presence of long-term humoral immunity. The only significant factor was disease severity. Patients with mild infections have more chance to have NAbs for a longer time. Age, gender, and comorbidity did not affect long-term NAb response. NAb titers decreased significantly over time, with an average rank of 24.0 versus 19.1 (p = 0.002). Multivariate generalized estimating equation analysis revealed that no parameter has an impact on the change of NAb titers over time. The majority of the late convalescent patients still had detectable low levels of neutralizing antibodies. The protective effect of these titers of NAbs from re-infections needs further studies.

SARS-CoV-2 Detection with De Novo-Designed Synthetic Riboregulators.

SARS-CoV-2 is a human pathogen and the main cause of COVID-19 disease, announced as a global pandemic by the World Health Organization. COVID-19 is characterized by severe conditions, and early diagnosis can make dramatic changes for both personal and public health. Low-cost, easy-to-use diagnostic capabilities can have a very critical role in controlling the transmission of the disease. Here, we are reporting a state-of-the-art diagnostic tool developed with an in vitro synthetic biology approach by employing engineered de novo riboregulators. Our design coupled with a home-made point-of-care device can detect and report the presence of SARS-CoV-2-specific genes. The presence of SARS-CoV-2-related genes triggers the translation of sfGFP mRNAs, resulting in a green fluorescence output. The approach proposed here has the potential of being a game changer in SARS-CoV-2 diagnostics by providing an easy-to-run, low-cost diagnostic capability.

Assessment of replication of bovine herpesvirus type 4 in human glioblastoma and breast cancer cells as a potential oncolytic virus.

Oncolytic viruses have been extensively used in cancer treatment due to their tropism, selective replication only in tumor cells, and possible synergic interaction with other therapeutics. Different researchers have demonstrated that bovine herpesvirus 4 (BoHV-4), a member of the gammaherpesviridae family, has oncolytic potential in some human-origin cancer cell lines like glioma through the selective replication strategy. Using four apoptosis detection methods, namely MTT, LDH, TUNEL, and Annexin V assays, we evaluated the apoptotic effect of BoHV-4 Movar33/63 reference strain along with a recombinant BoHV-4 expressing EGFP in U87 MG cells (human glioblastoma cell line), MDA MB-231 (human breast cancer cell line), and MCF10a (non-tumorigenic human mammary epithelial cell line). Our findings indicate that this virus can replicate and induce apoptosis in these cell lines and hinder in vitro proliferation in a dose-dependent manner. In conclusion, BoHV-4 has in vitro potential as a novel oncolytic virus in human cancer therapy. However, its replication potential in the MCF10a cells as a non-tumorigenic human mammary epithelial cell line is a concern in using this virus in cancer therapy, at least against human mammary tumors. Further studies must therefore be conducted to examine the specific apoptotic pathways induced by this virus to move on to further experiments.

Ribavirin shows antiviral activity against SARS-CoV-2 and downregulates the activity of TMPRSS2 and the expression of ACE2 in vitro.

Ribavirin is a guanosine analog with broad-spectrum antiviral activity against RNA viruses. Based on this, we aimed to show the anti-SARS-CoV-2 activity of this drug molecule via in vitro, in silico, and molecular techniques. Ribavirin showed antiviral activity in Vero E6 cells following SARS-CoV-2 infection, whereas the drug itself did not show any toxic effect over the concentration range tested. In silico analysis suggested that ribavirin has a broad-spectrum impact on SARS-CoV-2, acting at different viral proteins. According to the detailed molecular techniques, ribavirin was shown to decrease the expression of TMPRSS2 at both mRNA and protein levels 48 h after treatment. The suppressive effect of ribavirin in ACE2 protein expression was shown to be dependent on cell types. Finally, proteolytic activity assays showed that ribavirin also showed an inhibitory effect on the TMPRSS2 enzyme. Based on these results, we hypothesized that ribavirin may inhibit the expression of TMPRSS2 by modulating the formation of inhibitory G-quadruplex structures at the TMPRSS2 promoter. As a conclusion, ribavirin is a potential antiviral drug for the treatment against SARS-CoV-2, and it interferes with the effects of TMPRSS2 and ACE2 expression.

Phylogenetic analysis of West Nile virus: first report of lineage 1 in donkey in Turkey.

West Nile virus (WNV), a member of the Flaviviridae, is a major arbovirus that causes West Nile fever. Previous data showed the prevalence of the WNV serologically and molecular in Turkey, and the presence of lineage 1 in horses and humans has been reported. This is the first notification of partial phylogeny of WNV detected in donkeys in the northeast of Turkey (on the Iranian border). Blood serum samples collected from 25 donkeys without clinical symptoms were tested by RT-PCR. Sequence analysis of the sample detected as positive was performed. Multiple sequence alignments of reference sequences taken from GenBank were performed using the ClustalW method using the MEGA6 program. Partial nucleotide sequences of the capsid gene coding region revealed that the strains are closely related to viruses of lineage 1, clade 1a. According to the phylogenetic tree, the TUR/Igdir/donkey strain was included in the same cluster as the strain (KJ958922) previously obtained from horses in Turkey and the strain (GQ851658) from the Central African Republic. This study is the first report to show the circulation of WNV lineage 1 in donkeys in Turkey.

Akabane virus infection in Eastern Mediterranean Region in Turkey: Culicoides (Diptera: Ceratopogonidae) as a possible vector.

Akabane virus (AKAV), which causes Akabane disease, is an arthropod-borne virus (arbovirus) transmitted by Culicoides biting midges and mosquitoes. AKAV is an important pathogen that causes abortion and congenital anomalies in ruminants. In this study, we determined the prevalence of AKAV infection and identified possible viral vectors in Turkey's Eastern Mediterranean region. The presence and prevalence of AKAV infection were assessed using serological and virological methods. Serologically, the prevalence of AKAV antibodies in cattle, sheep and goats were 44.74% (400/894), 22.90% (60/262) and 14.52% (63/434), respectively, while the total prevalence was 32.89% (523/1590). AKAV-specific nucleic acid amplicons were obtained by real-time RT-PCR from 1.13% (9/799) and 1.74% (5/288) of the cattle and sheep tested, respectively. No goats were positive for AKAV RNA. Overall, AKAV-specific nucleic acid amplicons were detected in 0.87% (14/1604) of the sampled ruminants. In addition, specimens of the assumed vector, Culicoides, were caught using light traps and identified. Ten Culicoides species were detected in the area, of which Culicoides schultzei complex was the dominant species although 32 specimens could not be identified at the species level. These were defined as Culicoides spp. AKAV nucleic acid was detected in C. schultzei, Culicoides longipennis and Culicoides circumscriptus. Phylogenetic analysis indicated two different AKAV genogroups (genogroups Ib and genogroups II) while potential AKAV vectors in this region are C. schultzei complex, C. longipennis and C. circumscriptus.

Serological screening of West Nile virus among blood donors in northern Cyprus.

BACKGROUND: West Nile virus (WNV) is a neurotropic arbovirus that can also be transmitted through blood transfusion. Even though its geographic distribution has been expanding, there has not yet been any epidemiological data on WNV in northern Cyprus. The aim of our study is to fill this gap by using donated blood samples. METHODS: Samples collected from the main government hospital blood bank in Nicosia were analyzed by anti-WNV enzyme-linked immunosorbent assay (ELISA) (immunoglobulin M [IgM] and immunoglobulin G [IgG]). Seropositive samples were subjected to plaque reduction neutralization test (PRNT) for confirmation and analyzed by ELISA IgG avidity test and reverse transcription real-time polymerase chain reaction (rRT-PCR). RESULTS: Of the 760 sera samples, 2 (0.3%) were IgM+ and 31 (4.1%) were IgG+. Neutralization activity was detected in none (0.0%) of the IgM+ and 26 (83.9%) of IgG+ donor specimens. ELISA IgG avidity test reported high avidity in 21 (67.7%) and low avidity in one (3.2%) IgG+ sample. PRNT-confirmed anti-WNV IgG+ samples exhibited only borderline (19.2%) or high avidity (80.8%) values. rRT-PCR results were negative for both IgM+ and IgG+ samples. CONCLUSION: Anti-WNV antibodies were detected in northern Cyprus among blood donors. The establishment of preventive measures and evaluation of the geographic extent of the WNV in northern Cyprus are highly recommended.

Investigation of Crimean-Congo hemorrhagic fever virus in ruminant species slaughtered in several endemic provinces in Turkey.

A total of 1,337 serum and plasma specimens (939, 393 and 15 from cattle, sheep and goats, respectively) were collected monthly for one a year from ruminant species slaughtered in three Turkish cities endemic for Crimean-Congo hemorrhagic fever virus (CCHFV), Samsun, Sivas and Tokat. The serum samples were tested by commercial indirect ELISA to detect CCHFV antibodies, and positive or equivocal samples were later confirmed by a virus neutralization test (VNT). The seroprevalence in cattle, sheep, and goats was 36.21% (340/939), 6.27% (24/383), and 6.67% (1/15), respectively. Quantitative real-time RT-PCR was employed to detect viraemic animals at slaughter time. The percentage of CCHFV-viraemic animals was 0.67% (9/1337). The virus load varied between 4.1 x 101 and 2.4 x 103 RNA equivalent copies/mL in viraemic animals. The plasma samples that were positive for CCHFV genomic RNA were collected between April and May, when Hyalomma ticks are active. This study presents quantitative CCHFV load data in ruminant species at slaughter and interprets the likelihood of transmission for employees working in slaughterhouses in CCHFV-endemic regions.

Monitoring Crimean-Congo haemorrhagic fever virus RNA shedding in body secretions and serological status in hospitalised patients, Turkey, 2015.

IntroductionCrimean-Congo haemorrhagic fever (CCHF) is a tick-borne disease in Africa, Asia, the Balkan peninsula, the south-east of Europe and the Middle East, with mortality rates of 3-30%. Transmission can also occur through contact with infected animals or humans.AimThis observational, prospective case series aimed to investigate detectable viral genomic RNA in whole-body fluids and antibody dynamics in consecutive daily samples of patients diagnosed with CCHF until discharge from hospital.MethodsWe tested 18 patients and 824 swabs and sera with RT-PCR and 125 serum samples serologically.ResultsThe longest duration until clearance of viral RNA was 18 days from serum collection and 18, 15, 13, 19 and 17 days, respectively, from nasal, oral, genital (urethral or vaginal) and faecal swab, and urine. In seven patients, viral load decreased in serum at the same time as it increased in urine or persisted at the same logarithmic values. Despite clearance in serum, viral RNA was detected in faeces and genital swabs in two and three patients, respectively. Viral clearance from body fluids occurred earlier than from serum in eight patients on ribavirin treatment. The shortest seroconversion time was 3 days after symptom onset for IgM and IgG. Seroconversion of IgG occurred until Day 14 of symptoms.ConclusionWe report persistence of viral RNA in urine, faeces and genital swabs despite serum clearance. This may indicate a need for extending isolation precautions, re-evaluating discharge criteria and transmission risk after discharge, and considering oral swabs as a less invasive diagnostic alternative.

Molecular and antigenic characterization of bovine herpesvirus type 1 (BoHV-1) strains from cattle with diverse clinical cases in Turkey.

The present study reports the molecular and antigenic characterization of 13 bovine herpesvirus type 1 (BoHV-1) field viruses obtained from cattle with different clinical cases in Turkey between 1992 and 2017. We selected glycoprotein C (gC) of BoHV-1 as a target to detect and/or verify presence of the virus in suspect materials followed by virus isolation (VI) in MDBK cells. In seven out of 13 BoHV-1 positive samples, cytophatic effects (CPEs) were observed in MDBK cell cultures, although only four virus samples reached a sufficient titer to use in phylogenetic assay, restriction endonuclease analysis (REA), and virus neutralization test (VNT). According to the results of sequence analysis of the 13 BoHV-1 positive samples, nine BoHV-1 field viruses were determined as BoHV-1.1 and four as BoHV-1.2. Using REA, we demonstrated that two of our isolated viruses could be categorized as BoHV-1.1 while the other two isolates were BoHV-1.2 subtypes. Differences between the BoHV-1.1 and BoHV-1.2 isolates were also detected in the VNT results by assaying 125 suspected serum samples after testing with isolated (KY748023, KY748022, KY748020, and KY748021) and reference viruses (BoHV-1 Cooper and BoHV-5 Texas 89). These results are indicating the need to correctly identify BoHV-1 field isolates to better understand the epidemiology and pathogenesis of infection. In addition, it would be useful to identify the subtypes circulating in the specific geographical area while determining vaccination preferences.

Genetic variability human respiratory syncytial virus subgroups A and B in Turkey during six successive epidemic seasons, 2009-2015.

Human respiratory syncytial virus (HRSV) is most important viral respiratory pathogen of acute lower respiratory tract infections in infants and young children worldwide. The circulating pattern and genetic characteristics in the HRSV attachment glycoprotein gene were investigated in Turkey during six consecutive seasons from 2009 to 2015. HRSVA was dominant in the all epidemic seasons except 2011-2012 season. Partial sequences of the HVR2 region of the G gene of 479 HRSVA and 135 HRSVB were obtained. Most Turkish strains belonged to NA1, ON1, and BA9, which were the predominant genotypes circulating worldwide. Although three novel genotypes, TR-A, TR-BA1, and TR-BA2, were identified, they were not predominant. Clinical data were available for 69 HRSV-positive patients who were monitored due to acute lower respiratory tract illness. There were no significant differences in the clinical diagnosis, hospitalization rates, laboratory findings and treatment observed between the HRSVA and HRSVB groups, and co-infections in this study. The major population afflicted by HRSV infections included infants and children between 13 and 24 months of age. We detected that the CB1, GB5, and THB strains clustered in the same branch with a bootstrap value of 100%. CB-B and BA12 strains clustered in the same branch with a bootstrap value of 65%. The BA11 genotype was clustered in the BA9 genotype in our study. The present study may contribute on the molecular epidemiology of HRSV in Turkey and provide data for HRSV strains circulating in local communities and other regions worldwide.

MicroRNA expression profiles in patients with acute Crimean Congo hemorrhagic fever reveal possible adjustments to cellular pathways.

Several viral diseases are associated with altered microRNA (miRNA) expression, which can provide vital information about how cellular pathways respond to infection. However, the miRNA profile of Crimean Congo Hemorrhagic Fever (CCHFV) infections are not known. To address this gap, we performed real-time PCR-based miRNA analysis in individuals with acute Crimean Congo Hemorrhagic Fever (CCHFV) infections, with the goal of identifying pathways that might be associated with this disease. Peripheral blood mononuclear cells were analysed in eight individuals with detectable viral RNA and compared to five healthy subjects. A total of 106 differentially expressed miRNAs were identified, of which 19 miRNAs were either fivefold prominently up- or down-regulation. Several miRNAs associated with cytokine expression, some of which were previously associated with Dengue and Hantavirus infections were revealed. Moreover, possible mechanisms related to secretion of adhesion molecules and viral escape from innate immunity were identified. Pathway enrichment analyses further revealed the putative involvement of TNF-alfa, TGF-beta, MAPK, WNT, and neurotrophin signaling pathways in disease pathogenesis. J. Med. Virol. 89:417-422, 2017. © 2016 Wiley Periodicals, Inc.

[Multicenter investigation of bufavirus in the etiology of viral central nervous system infections of adults and children]. / Bufavirusun çocuklar ve eriskinlerdeki viral santral sinir sistemi enfeksiyonlarinin etiyolojisi açisindan çok Mmerkezli olarak arastirilmasi.

Bufavirus (BuV) is a newly-identified parvovirus in the family of Parvoviridae. Metagenomic analysis of fecal samples from children in Burkina Faso with acute diarrhea showed a highly divergent parvovirus, which was named bufavirus (BuV). The global distribution, epidemiology and genetic characteristics of BuVs infections are obscure. It was first discovered as an agent causing gastroenteritis but the association of BuV infections with various clinical presentations mostly remain to be explored. The aims of this study were to investigate probable impact of BuV in central nervous system infections in a region where it was previously reported to cause human infections and to detect enteroviruses (EV) which are reported as a cause of central nervous system infections in our country. The study was undertaken in three institutions in Ankara province, Central Anatolia, Turkey. Patients, clinically diagnosed with febrile disease and/or central nervous system infections of presumed viral etiology, were enrolled in the study with informed consent. Cerebrospinal fluid specimens were collected from 93 children attended to Gazi University Hospital and Diskapi Yildirim Beyazit Hospital from October 2011-April 2015 and 33 adult patients, attended to Hacettepe University Hospital from June 2012 to March 2013. Clinical history and follow-up, physical examination and standard laboratory findings of the patients were recorded. Nucleic acid extraction was performed via commercially available spin-column assays and complementery DNA (cDNA) synthesis was performed by using commercially available cDNA synthesis kit with randomised hexamer primers. BuV detection was carried out by in house nested-polymerase chain reaction (PCR) utilized with previously-described primers. EV detection was carried out by in house PCR with pan-enterovirus primers. Seventy-four percent (93/126) and 26% (33/126) of the patients were children (0-18) and adults (19-86), respectively. In all patients, bacterial, mycobacterial and fungal cultures were negative, as well as PCR for herpes simplex virus (HSV) types 1 and 2. PCR results of all samples were negative for BuV and EV. This is the first study that evaluates a probable association of BuV and central nervous system infections. Although Parvovirus B19, a well-characterized human pathogen can rarely cause encephalitis, our findings did not confirm such an association for BuV in this preliminary investigation. However, long-term evaluation of individual cases with unknown etiology is required to reveal the relationship of the virus with specific environments.

Biosafety standards for working with Crimean-Congo hemorrhagic fever virus.

In countries from which Crimean-Congo haemorrhagic fever (CCHF) is absent, the causative virus, CCHF virus (CCHFV), is classified as a hazard group 4 agent and handled in containment level (CL)-4. In contrast, most endemic countries out of necessity have had to perform diagnostic tests under biosafety level (BSL)-2 or -3 conditions. In particular, Turkey and several of the Balkan countries have safely processed more than 100 000 samples over many years in BSL-2 laboratories. It is therefore advocated that biosafety requirements for CCHF diagnostic procedures should be revised, to allow the tests required to be performed under enhanced BSL-2 conditions with appropriate biosafety laboratory equipment and personal protective equipment used according to standardized protocols in the countries affected. Downgrading of CCHFV research work from CL-4, BSL-4 to CL-3, BSL-3 should also be considered.

[Determination of cytomegalovirus glycoprotein B genotypes in different geographical regions and different patient groups in Turkey]. / Türkiye'de farkli cografi bölgelerde ve farkli hasta gruplarindaki sitomegalovirus izolatlarinin glikoprotein B genotiplerinin belirlenmesi.

Cytomegalovirus (CMV), a common virus found all around the world, usually causes asymptomatic infections in immunocompetent hosts, however it may lead to serious complications in immunodeficient patients and in the fetus. CMV is divided into four genotypes according to the polymorphisms in UL55 gene that encodes for envelope glycoprotein B. Nucleotide polymorphisms of CMV gB gene can affect the cell tropism of the virus and host immune response and believed to have important changes in the pathogenesis of CMV. The aim of this study was to determine the gB genotypes of CMV isolates from different patient groups selected from different regions of Turkey. A total of 136 clinical specimens from patients (66 female, 70 male; age range: 0-65 years, mean age: 24.03 ± 17.17) who were diagnosed to have CMV infection by polymerase chain reaction (PCR) and/or antigenemia tests, between 2001-2014, in the medical school hospitals of Akdeniz, Ege, Istanbul Cerrahpasa and Erciyes Universities (located at Mediterranean, Aegean, northwest and central Anatolia regions, respectively), were included in the study. The patient group consisted of 80 renal transplant (RT) recipients, 35 stem cell transplant (SCT) recipients, 13 newborns, seven heart transplant (HT) recipients and one pregnant woman. CMV gB genotypes were determined by PCR-RFLP (restriction fragment length polymorphism) method, and DNA sequencing and phylogenetic analysis were performed for the randomly selected 15 isolates with different genotypes. Among 136 (135 plasma, 1 amnion fluid) samples, the most frequent genotype was gB1 (n= 44, 32.4%), followed by gB2 (n= 39, 28.6%), gB3 (n= 36, 26.5%) and gB4 (n= 8, 5.9%); however nine (6.6%) samples could not be genotyped. When analysis were interpreted according to the patient groups, it was determined that the genotypes in RT recipients were gB1 32.3%, gB2 28.7%, gB3 26.5% and gB4 5.9%; in SCT recipients gB1 34.3%, gB2 28.6%, gB3 22.9% and gB4 5.7%; in HT recipients gB3 57.1%, gB1 14.3% and gB2 14.3%; in newborns gB1 38.4%, gB3 30.8%, gB2 15.4% and gB4 7.7%, and gB2 genotype in the pregnant woman. As our study was a descriptive study to determine the genotypes of CMV gB, the relationship between the genotypes and the variants such as viral load, symptomatic disease and prognosis were not analyzed. As a result, the isolation of different gB genotypes in various case groups from four distinctive provinces, underlines the diversity of CMV gB genotypes in Turkey.

Close relationship between West Nile virus from Turkey and lineage 1 strain from Central African Republic.

We sequenced West Nile viruses (WNVs) from Turkey and found close relationships to WNV lineage 1 strain ArB310/67 from the Central African Republic, distinct from other WNVs circulating in the Mediterranean Basin, eastern Europe, and the Middle East. These findings suggest independent introductions of WNV strains from Africa to the Middle East.

Prospective investigation of the impact of West Nile Virus infections in renal diseases.

An increased incidence of chronic kidney disease (CKD) after West Nile Virus (WNV) infections has been suggested but the association of WNV infections with renal damage remain inconclusive. This study was undertaken to characterize WNV infections in individuals with acute kidney injury (AKI) and CKD, and to evaluate hemodialysis as a probable transmission route. A total of 463 plasma and urine samples were collected from 45 AKI and 77 CKD patients. Nested and real-time polymerase chain reaction (PCR) assays were employed for viral RNA detection. Specific immunoglobulins were investigated via immunofluorescence and plaque reduction neutralization assays. Consecutive pre and post-dialysis samples were evaluated in CKD cases. WNV RNA and specific immunoglobulins were detected in 7 (5.7%) and 5 (4.1%) individuals, respectively. The AKI patients with WNV RNA in blood and urine had underlying diseases requiring immunosuppressive therapy and demonstrated moderate to high viral loads. No clinical symptom related to WNV infection were observed in CKD cases with detectable viral nucleic acids. All WNV sequences were characterized as lineage 1 clade 1a and several amino acid substitutions with unknown impact were noted. Detailed epidemiologic investigation of WNV RNA positive CKD cases revealed probable vector-borne virus exposure, without the evidence for transmission via hemodialysis.