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The serious diseases of the central nervous system (CNS); encephalitis and meningitis, have high mortality and morbidity rate especially not diagnosed and treated in time. Nucleic acid testing (NAT) is the tool of choice for viral diagnosis in CNS infections. In this study, viral etiological agents found in cerebrospinal fluid (CSF) samples sent to our university hospital virology laboratory for laboratory diagnosis of CNS infections were retrospectively evaluated and results were compared with other reports from our country. Viral etiological agents found in cerebrospinal fluid (CSF) samples sent to Ege University Faculty of Medicine Department of Medical Microbiology Virology Laboratories for laboratory diagnosis of CNS infection between 01.01.2009-31.12.2015 were evaluated retrospectively. A total of 3778 CSF tests were performed for cell culture of enterovirus (EV) in 487 samples and 3291 tests for nucleic acid testing (NAT) by real time polymerase chain reaction (PCR) in herpes simplex virus 1 (HSV1), herpes simplex virus 2 (HSV2), varicella zoster virus (VZV), Epstein-Barr virus (EBV), cytomegalovirus (CMV), human herpes virus 6 (HHV6) and EV. VZV and EV NAT's were performed during the last one and five years period, respectively. NAT positive results for HSV1, HSV2, CMV, EBV, VZV, HHV6 and EV were 1.80% (24/1333), 0.08% (1/1333), 3.28% (19/580), 4.35% (22/506), 0.46% (1/216), 1.05% (5/478) and 3.37% (6/178), respectively. EV was isolated in 30 (6.20%) of 487 CSF samples by viral culture. Positive samples were mainly from pediatric, neurology and infectious diseases clinics as expected. The number of higher positive results were found in samples sentin december (35.3%), july (12.9%) and november (10.6%). Overall 80% of positive samples belonged to patients over 18 years old. When the results of other studies reported from Turkey are examined, although the positivity rates are generally similar, it is seen that the rates specific to certain factors are higher in selected smaller patient groups like HSV1 and EV. Rapid nucleic acid tests like multiplex PCR and microarray will provide more practical and effective laboratory diagnosis approach in CNS infections, since many more microorganisms may be causative agents.

Herpes simplex virus (HSV) infections are a common clinical problem worldwide. HSV infections have a severe and rapidly progressive course especially in immunocompromised patients, leading to significant morbidity and mortality. Therefore, rapid and reliable laboratory diagnosis of HSV infections is of crucial importance for the initiation of early antiviral therapy. In this study the aim was to investigate the presence of HSV type 1 and type 2 in clinical specimens by cell culture, in-house polymerase chain reaction (PCR) and direct fluorescein antibody (DFA) methods. The study was conducted at Erciyes University Gevher Nesibe Hospitals between March 2006 and june 2007. A total of 65 clinical specimens, 38 of them being cerebrospinal fluid (CSF) samples obtained from meningoencephalitis suspected cases and 27 being vesicular/conjunctival/genital swabs obtained from patients with different clinical presentations (13 herpes labialis, 5 keratoconjunctivitis, 4 gingivostomatitis, 3 eczama herpeticum, 1 herpetic whitlow, 1 genital ulcer). The age range of the 65 patients varied between 1-68 years, 20 being children. All the samples except CSF were investigated by 3 of the test methods, however, CSF samples were tested only by cell culture and PCR since DFA is not recommended. HSV was found positive in 48.1% (13/27) of the 27 swab specimens by DFA, in 66.6% (18/27) by PCR and in 51.8% (14/27) cytopathic effect consistent with HSV was observed. HSV positivity was detected in 7.8% (3/38) of the 38 CSF specimens by cell culture and in 47.4% (18/38) by in-house PCR. Viral growth in cell cultures showing cytopathic effect were further confirmed by DFA method using HSV-1 and HSV-2 specific fluorescein monoclonal antibodies (Monofluo, Bio-Rad Laboratories, USA). Agreement between the methods were investigated by Kappa analysis. Moderate level agreement was determined for both swab (K = 0.7) and CSF (K = 0.6) specimens for the tested methods when cell culture was considered as the reference method. In conclusion for swab specimens, primarily DFA which is a practical and rapid method, could be applied. This step might be followed by PCR and cell culture techniques sequentially. On the other hand CSF specimens should be investigated by the rapid and more sensitive PCR method.

Herpes simplex virus (HSV) meningoencephalitis has a high mortality rate if proper antiviral therapy is not applied. Thus rapid diagnosis is of peculiar importance in such cases. In this study we aimed to evaluate the use of polymerase chain reaction (PCR) and detection of intrathecally synthesized antibodies by serological methods in viral meningoencephalitis suspected cases to determine HSV as the causative agent. Seventeen cases with cerebrospinal fluid (CSF) samples with microscopical and biochemical findings compatible with viral encephalitis were included in this study. CSF samples yielded no bacterial growth. Cell cultures propagated in Vero cell line and PCR with different primer sets against HSV-1 and HSV-2 (specific for US7 and US2 gene regions, respectively) were used to investigate the presence of HSV in CSF. Serum samples were taken simultaneously with the CSF sampling and "Reibergram" graphics and antibody index (AI) calculations were used for the evaluation of intrathecal antibody synthesis. Albumin and total IgG levels in serum and CSF samples measured with nephelometry (Dade-Behring, Germany) for Reibergram graphics, albumin and IgG ratios were calculated. Quantitative levels of HSV 1+2 IgG were measured in serum and CSF samples using ELISA (HSV-1/2 Pool; Antibody Determination in CSF, Euroimmun, Germany) for AI determination. One of the CSF samples obtained one week after the neurological symptoms had started, yielded HSV-2 in cell culture and also HSV-2 DNA was detected by PCR. Intrathecal antibody synthesis was not detected in this case. In two cases with symptoms lasting for more than three weeks, intrathecal IgG synthesis in Reibergrams and pathological intrathecal HSV AI (27.9 and 7.9, respectively) were detected, however, virus isolation and PCR detection were not successful. For the other 14 cases, HSV-DNA were found negative and no intrathecal antibody synthesis were detected. HSV meningoencephalitis can be diagnosed via using PCR which has been accepted as gold standard in recent years, with 24 hours turn-around time. However, if CSF samples were taken later than the first week following the beginning of symptoms, possibility of HSV detection by PCR is lowered. According to the data obtained from this limited study, it may be suggested that PCR may be used for the detection of HSV-DNA in CSF samples during the early phase of meningoencephalitis cases, however, consideration must be taken to detect the intrathecal antibodies during the later phases of the infection where intrathecal antibody synthesis starts.

Acute otitis media with effusion (OME) is one of the major causes of antibiotic use, indication for operation and hearing loss in children. In two third of the cases the etiologic agents are bacteria. Nonetheless, increasing numbers of reports have implicated viruses as etiologic agents that may have some effect on prognosis of OME. The aim of this study was to investigate the presence of nucleic acids of respiratory syncytial virus (RSV) type A and B, influenza type A virus, adenovirus, cytomegalovirus (CMV), herpes simplex virus type-1 (HSV-1), and enteroviruses in the middle ear effusion specimens from children with otitis media by TaqMan real-time PCR. As a result, 18 of 30 (60%) OME samples were found positive in terms of viral nucleic acids by real-time PCR. RSV-A was detected in nine samples (30%), CMV in 3 (10%) samples and HSV-1 in 1 (3.3%) sample. In five of the samples two viruses were detected in the same sample (three were positive for adenovirus and RSV-A, and two were positive for CMV and RSV-A). Our data have supported the importance of viruses as etiologic agents of OME. Additionally, it was thought that TaqMan real-time PCR may be used as a reliable and rapid method for the detection of viruses in the middle ear effusion samples.

Mutations or variants that impair function of ribonuclease L (RNase-L), particularly R462Q, have been proposed as susceptibility factors for the innate antiviral response. The aim of this study was to investigate and compare the expression levels of RNase-L and mutation of R462Q in the tonsils of tonsillectomy patients who were infected and not infected with herpes simplex virus type 1 (HSV-1), Epstein-Barr virus (EBV) and human herpes virus 6 (HHV-6). Six tonsils were included in the study. One tonsil was infected with all of these three viruses, two were infected with at least one of these viruses, and three were not infected with these viruses. The presence of viral DNAs in the tonsil tissues had been searched by polymerase chain reaction (PCR) in our previous study. Reverse transcriptase PCR method was used for RNase-L expression analyses, and single strand conformation polymorphism (SSCP) and direct sequencing methods were used for the mutation analyses. PCR products containing R462Q mutation site in the genomic DNA were used for SSCP analysis. In addition to SSCP analyses, partial sequencing of the cDNA PCR product containing R462Q mutation site were performed. As a result, no difference between the virus-infected and non-infected tonsils for the expressions of RNase-L were detected, and there were no mutations detected by SSCP and sequencing analyses. It was concluded that other factors than RNase-L protein, might be involved in the innate defense mechanisms of tonsil cells against viruses.

In this report, characteristics of two cases of Herpes simplex virus (HSV) encephalitis with normal cerebrospinal fluid (CSF) findings at the time of admission have been discussed and the current literature has been reviewed. The diagnosis of the cases (one was 23 years old male, and the other was 75 years old female patient) was made on the magnetic resonance imaging (MRI) findings concordant with HSV encephalitis, together with HSV-1 DNA positivity by polymerase chain reaction (PCR). Both of the patients were treated with acyclovir (3 x 750 mg/day) lasting for 15 days and 21 days, respectively. The first male patient recovered with mild neurological defects, whereas the second female patient died because of nosocomial pneumonia and septicemia. In conclusion, even the CSF findings are normal, in cases considered to be HSV encephalitis, MRI should be the first radiological diagnostic step and the diagnosis should be confirmed by the detection of HSV DNA in CSF by PCR.

The aim of this study was to investigate the presence of Herpes simplex virus (HSV) type 1 and type 2 DNA from the clinical samples sent to our routine laboratory, by real-time polymerase chain reaction (PCR). A total of 328 samples collected from 306 female and 7 male patients who were admitted to different outpatient clinics were included in the study. The samples included 235 cervical swab samples (of which 150 were from pregnant women), 77 amniotic fluid, 8 blood, 6 cerebrospinal fluid (CSF), one pericardial fluid and one cervical biopsy. DNA extraction were performed with High Pure Viral Nucleic Acid Kit (Roche, Germany) and amplified in Light Cycler (Roche, Germany) with a commercial amplification mix (Metis Biotechnology, Ankara). HSV-DNA positivity were found in 2.1% of the cervical samples (three of 150 pregnant and two of 85 non-pregnant women), two of the blood samples and one of the CSF sample, while there were no positive result for the other clinical specimens. It can be concluded that, real-time PCR would be preferred in conditions requiring rapid diagnosis such as HSV infections of central nervous system and HSV suspected infections of immunosupressed patients, as a rapid and practical method.

The aim of this study was to investigate the presence of Herpes simplex virus type 1, in the specimens from patients clinically prediagnosed as herpetic keratitis or keratoconjunctivitis, by virus isolation and direct immunoperoxidase methods. The samples obtained from a total of 33 patients included ulcerated corneal epithelium scrapings and, swabs from ulcer, corneal epithelium, and lower conjunctivas. For virus isolation, both scraping and swab samples for each patient, were inoculated into monolayered Vero cell lines which have previously cultivated on 24-well plates, and examined for the presence of cytopathic effects typical for HSV-1. At the end of 5-days incubation period, the culture media were discarded, the cells were fixed and stained with peroxidase labeled specific HSV-1 antibodies (direct immunoperoxidase--DIP--method). Simultaneously, the smears were prepared from the samples which were sufficient in amount, and stained with DIP method. As a result, cytopathic effects on cell cultures were observed in 4 (12.1%) of the samples, of which 2 were also positive with DIP method. Following DIP staining of smears prepared from 15 samples, HSV-1 antigen positivity was detected in only 1 of the samples. This sample was negative in cell culture. In contrast, one of the 2 cell culture positive samples yielded negative result in smear examination, while the other could not be examined since the sample was not enough. In conclusion, HSV-1 has been shown as the etiologic agent in 3 (9.1%) of our patients clinically prediagnosed as herpetic keratitis or keratoconjunctivitis, and lesion scrapings were determined as the most appropriate samples for virus isolation.