The assessment of microbial infection in children with autism spectrum disorders and genetic folate cycle deficiency.

BACKGROUND: The results of disparate clinical studies indicate abnormally frequent cases of certain microorganisms in children with autism spectrum disorders (ASD). However, these data require clarification and systematization. The study aims to study the structure of the microbial profile in children with ASD and genetic folate cycle deficiency (GFCD) and consider differences in diagnostic approaches for identifying microorganisms of different types. METHODS: The study analyzed medical data from 240 children (187 boys and 63 girls) with GFCD aged 2 to 9 years. The children had clinical manifestations of ASD (the study group, SG). The control group (CG) included 53 clinically healthy children (37 boys and 16 girls) of the same age but without GFCD. Both groups of children were tested on active herpetic infections (HSV-1/2, VZV, EBV, CMV, HHV-6, HHV-7, HHV-8), ТТV, Streptococcus pyogenes, Candida albicans, Borrelia burgdorferi, Mycoplasma pneumoniae, Chlamydia pneumoniae, Yersinia enterocolitica, Toxoplasma gondii, congenital CMV neuroinfection and postnatal HSV-1/2 encephalitis. The testing used diagnostic methods specified in PubMed-indexed studies. RESULTS: In the SG, TTV was found in 196 children (82%), HHV-7 - in 172 (72%), HHV-6 - in 162 (68%), EBV - in 153 (64%), Streptococcus pyogenes - in 127 (53%), Candida albicans - in 116 (48%), Borrelia - in 107 (45%), Mycoplasma pneumoniae - in 94 (39%), Chlamydia pneumoniae - in 85 (35%), Yersinia entеrocolitica - in 71 (30%), Toxoplasma gondii - in 54 (23%), congenital CMV neuroinfection - in 26 (11%), and postnatal HSV-1/2 encephalitis - in 11 children (5% of cases) (p < p0.05; Z < Z0.05). In the SG, there was a higher microbial load in older children (p < p0.05; Z < Z0.05). No gender differences were found. CONCLUSIONS: The study described and characterized a specific abnormal microbial spectrum with a predominance of viral opportunistic agents in children with ASD associated with GFCD.

Research Note: Simultaneous detection of infectious laryngotracheitis virus, fowlpox virus, and reticuloendotheliosis virus in chicken specimens.

Infectious laryngotracheitis (ILT), fowlpox (FP), and reticuloendotheliosis are important poultry diseases caused by gallid herpesvirus 1 (ILTV), fowlpox virus (FWPV), and reticuloendotheliosis virus (REV), respectively. Coinfections with ILTV and FWPV occur naturally in chickens, and FP in its more virulent wet form is characterized by diphtheritic lesions and easily confused with ILT. Moreover, the insertion of only partial REV-LTR or a nearly full-length REV into the FWPV genome, located between the ORF 201 and ORF 203, has increased recently in wild-type field FWPV isolates. Therefore, it is critical to detect ILTV, FWPV, REV-integrated FWPV, and REV early and accurately. In this study, we successfully developed a multiplex PCR assay for the simultaneous detection of ILTV, FWPV, REV-integrated FWPV, and REV, and the detection limits was 1 × 54 copies/tube. When used to test clinical samples, the results of the multiplex PCR were in 100% agreement with singleplex PCRs and sequencing. This new multiplex PCR is a simple, rapid, sensitive, specific, and cost-effective method for detection of 4 viruses in clinical specimens.

Increased incidence of systemic serious viral infections in patients with inflammatory bowel disease associates with active disease and use of thiopurines.

Background: The magnitude and drivers of the risk of serious viral infections in Inflammatory Bowel diseases (IBD) are unclear. Objective: The objective of this study was to assess the incidence and risk factors for systemic serious viral infections in IBD patients. Methods: Using MICISTA, a database detailing prospective characteristics and complications of IBD, we identified patients that were followed for IBD in 2005-2014 outside the context of organ transplantation, HIV infection or chronic viral hepatitis. We estimated incidences of systemic serious viral infections, defined by the need for hospitalization or permanent organ damage. Standardized incidence ratios (SIRs) were calculated using the French hospital database. We performed a case-control study nested in MICISTA for assessing the role of exposure to IBD drugs and IBD clinical activity in the risk of developing infection. Results: We identified 31 patients with serious viral infections among 2645 patients followed for 15,383 person-years. We observed 13 cases of cytomegalovirus, 10 Epstein-Barr virus, 5 varicella zoster virus and 3 herpes simplex virus infections. No deaths occurred. The incidence rate of infections in patients with IBD was 2.02/1000 person-years, and the SIR was 3.09 (95% confidence interval (CI), 1.98-4.20; p = 0.0002) in the study population. By multivariate analysis, increased risk of infection was associated with exposure to thiopurines (odds ratio (OR), 3.48; 95% CI, 1.36-8.90; p = 0.009), and clinically active IBD at onset of infection (OR, 3.35; 95% CI, 1.23-9.23; p = 0.02). Conclusions: The incidence of systemic serious viral infections in patients with IBD is tripled compared to general population. Clinically active IBD and exposure to thiopurines are the main drivers of the risk.

Clinical Results and Utility of Herpesviruses Multiplex Polymerase Chain Reaction: Assessment of Aqueous Humor Samples From Patients With Corneal Endotheliitis and High Intraocular Pressure.

PURPOSE: The main purpose of this study was to evaluate herpesvirus infection in patients with corneal endotheliitis and high intraocular pressure (IOP) using multiplex polymerase chain reaction (PCR) in aqueous humor samples. MATERIALS AND METHODS: This was a retrospective, observational study of immunocompetent patients living in South Korea. Eligible subjects had typical corneal endotheliitis with an IOP≥21 mm Hg or required antiglaucoma medication. Multiplex PCR was performed using aqueous humor samples obtained at first visit to detect the DNA of 6 herpesviruses. RESULTS: Forty-two eyes from 42 patients with >6 months' follow-up were analyzed. Of these, 16 were herpesvirus-positive: 3 herpes simplex virus 1, 3 varicella-zoster virus, 9 cytomegalovirus, and 1 Epstein-Barr virus. Eyes with coin-shaped or fine keratic precipitates (kps), high IOP, and a low baseline endothelial cell count were more likely to show a positive result on multiplex PCR. Univariate analysis showed that male sex (P=0.014), a previous history of uveitic glaucoma (P=0.048), and the presence of fine kps (P=0.031) were significantly associated with a positive PCR result. On multivariate analysis, male sex (P=0.010) and a previous history of uveitic glaucoma (P=0.031) showed a significant positive association. CONCLUSIONS: Cytomegalovirus was the most commonly detected herpesvirus in patients with corneal endotheliitis and high IOP. A positive PCR result was seen more frequently in male individuals and patients with fine kps or a history of uveitic glaucoma.

Detection of Herpesvirus anguillae (AngHV-1) in European eel Anguilla anguilla (L.) originating from northern Poland-assessment of suitability of selected diagnostic methods.

The Community Action Plan requests EU member states to implement measures that ensure the recovery of the severely depleted European eel stocks. One of the main threats is posed by Anguillid herpesvirus 1 (AngHV-1) leading to increased mortality in both wild and farmed eels. Following recommendations of the OIE to minimize the risk of obtaining false-negative results, the main aim of the study was to optimize diagnostic methods for AngHV-1 detection using conventional PCR, nested PCR and in situ hybridization assay. While 53.3% of the individual organ samples were tested positive for AngHV-1 by PCR, the additional virus analysis via nested PCR revealed that the actual prevalence was 93.3%. In the cell cultivation passages, a cytopathic effect was hardly found in the first two rounds. In the third passage onto cell cultures, a lytic CPE was detected. The identification and confirmation of the viruses obtained from cell cultures as well as directly from the organ tissues were proceeded by PCR, nested PCR and sequencing of the PCR products. While no positive signal was detectable in the first round by PCR using samples from the third cell culture passages, the nested PCR provided weak but visible positive signals.

Assessment of quantitative polymerase chain reaction for equine herpesvirus-5 in blood, nasal secretions and bronchoalveolar lavage fluid for the laboratory diagnosis of equine multinodular pulmonary fibrosis.

REASONS FOR PERFORMING STUDY: The ante mortem diagnosis of equine multinodular pulmonary fibrosis (EMPF) relies on histopathological results and polymerase chain reaction (PCR)-positive equine herpesvirus (EHV)-5 testing of lung tissue. Polymerase chain reaction detection of EHV-5 in bronchoalveolar lavage fluid (BALF) is commonly used to support a diagnosis of EMPF. However, the diagnostic power of EHV-5 testing on BALF and other biological samples such as blood and nasal secretions has yet to be shown to support a diagnosis of EMPF. OBJECTIVES: To determine the frequency of detection and the viral loads of EHV-5 by quantitative PCR (qPCR) in blood, nasal secretions and BALF from horses confirmed with EMPF, healthy horses and horses with non-EMPF pulmonary diseases. STUDY DESIGN: Prospective study. METHODS: The study population consisted of 70 adult horses divided into 4 groups based on a combination of clinical findings, cytology of BALF, imaging studies of the thoracic cavity and histopathology of pulmonary tissue: control group (n = 14), EMPF group (n = 11); inflammatory airway disease group (n = 32); and non-EMPF interstitial lung disease group (n = 13). For each horse, whole blood, nasal secretions and BALF were available for EHV-5 qPCR testing. Sensitivities, specificities and their respective 95% confidence intervals were calculated for viral loads from blood, nasal secretions and BALF. In addition, these measures were calculated for combined use of blood and nasal secretions. RESULTS: The detection of EHV-5 in BALF was strongly associated with EMPF (sensitivity 91%, specificity 98.3%). Detection of EHV-5 in blood was, independent of the viral loads, strongly associated with EMPF with a sensitivity of 91% and specificity of 83.1%. The detection of EHV-5 in nasal secretions displayed the highest sensitivity (72.7%) and specificity (83.1%) at a level of >245,890 glycoprotein B target genes/million cells to support a diagnosis of EMPF. Dually positive blood and nasal secretions at any viral loads in support of EMPF yielded a sensitivity and specificity of 90% and 89.8%, respectively. CONCLUSIONS: Although histopathological confirmation (lung biopsy) is considered the gold standard for EMPF diagnosis, results of qPCR testing of BALF or a combination of whole blood and nasal secretions should be regarded as clinically useful in support of this diagnosis. The latter testing may be relevant when dealing with horses in respiratory distress, for which invasive procedures such as BALF collection or lung biopsies may be detrimental to their health.

Translation of a laboratory-validated equine herpesvirus-1 specific real-time PCR assay into an insulated isothermal polymerase chain reaction (iiPCR) assay for point-of-need diagnosis using POCKIT™ nucleic acid analyzer.

Equine herpesvirus myeloencephalopathy (EHM), a major problem for the equine industry in the United States, is caused by equine herpesvirus-1 (EHV-1). In addition, EHV-1 is associated with upper respiratory disease, abortion, and chorioretinal lesions in horses. Here we describe the development and evaluation of an inexpensive, user-friendly insulated isothermal PCR (iiPCR) method targeting open reading 30 (ORF30) to detect both neuropathogenic and non-neuropathogenic strains on the field-deployable POCKIT™ device for point-of-need detection of EHV-1. The analytical sensitivity of the EHV-1 iiPCR assay was 13 genome equivalents per reaction. The assay did not cross react with ten non-target equine viral pathogens. Performance of the EHV-1 iiPCR assay was compared to two previously described real-time PCR (qPCR) assays in two laboratories by using 104 archived clinical samples. All 53 qPCR-positive and 46 of the 51 qPCR-negative samples tested positive and negative, respectively, by the iiPCR. The agreement between the two assays was 95.19% (confidence interval 90.48-99.90%) with a kappa value of 0.90. In conclusion, the newly developed EHV-1 iiPCR assay is robust to provide specificity and sensitivity comparable to qPCR assays for the detection of EHV-1 nucleic acid in clinical specimens.

A potentiometric biosensor for rapid on-site disease diagnostics.

Quantitative point-of-care (POC) devices are the next generation for serological disease diagnosis. Whilst pathogen serology is typically performed by centralized laboratories using Enzyme-Linked ImmunoSorbent Assay (ELISA), faster on-site diagnosis would infer improved disease management and treatment decisions. Using the model pathogen Bovine Herpes Virus-1 (BHV-1) this study employs an extended-gate field-effect transistor (FET) for direct potentiometric serological diagnosis. BHV-1 is a major viral pathogen of Bovine Respiratory Disease (BRD), the leading cause of economic loss ($2 billion annually in the US only) to the cattle and dairy industry. To demonstrate the sensor capabilities as a diagnostic tool, BHV-1 viral protein gE was expressed and immobilized on the sensor surface to serve as a capture antigen for a BHV-1-specific antibody (anti-gE), produced in cattle in response to viral infection. The gE-coated immunosensor was shown to be highly sensitive and selective to anti-gE present in commercially available anti-BHV-1 antiserum and in real serum samples from cattle with results being in excellent agreement with Surface Plasmon Resonance (SPR) and ELISA. The FET sensor is significantly faster than ELISA (<10 min), a crucial factor for successful disease intervention. This sensor technology is versatile, amenable to multiplexing, easily integrated to POC devices, and has the potential to impact a wide range of human and animal diseases.

Homogenization with heat treatment: A cost effective alternative to nucleic acid extraction for herpes simplex virus real-time PCR from viral swabs.

Most laboratories use expensive commercial kits to purify nucleic acids and remove PCR inhibitors that may be present in clinical specimens. In this study a simple homogenization with heat treatment of herpes simplex virus types 1 and 2 (HSV-1/2) was shown to be equivalent to commercial kit-based nucleic acid extraction methods. With a cost of less than $1 USD per extraction, this method provides an economical, rapid, and effective method to recover HSV-1/2 DNA from swabs suitable for real-time HSV PCR.

Viral infections affecting the skin in organ transplant recipients: epidemiology and current management strategies.

Viral skin infections are common findings in organ transplant recipients. The most important etiological agents are the group of human herpesviruses (HHV), human papillomaviruses (HPV), and molluscum contagiosum virus. HHV that are important in this group of patients are herpes simplex virus (HSV) types 1 and 2, varicella-zoster virus (VZV), cytomegalovirus (CMV), Epstein-Barr virus (EBV), HHV-6 and -7, and HHV-8, which causes Kaposi sarcoma (KS). HSV infections are characterized by their ability to establish latency and then reactivate at a later date. The most common manifestations of HSV infection in organ transplant recipients are mucocutaneous lesions of the oropharynx or genital regions. Treatment is usually with acyclovir, valaciclovir, or famciclovir. Acyclovir resistance may arise although the majority of acyclovir-resistant strains have been isolated from AIDS patients and not organ transplant recipients. In such cases, alternatives such as foscarnet, cidofovir, or trifluridine may have to be considered. VZV causes chickenpox as well as herpes zoster. In organ transplant recipients, recurrent herpes zoster can occur. Acute chickenpox in organ transplant patients should be treated with intravenous acyclovir. CMV infection occurs in 20-60% of all transplant recipients. Cutaneous manifestations, which include nonspecific macular rashes, ulcers, purpuric eruptions, and vesiculobullous lesions, are seen in 10-20% of patients with systemic infection and signify a poor prognosis. The present gold standard for treatment is ganciclovir, but newer drugs such as valganciclovir appear promising. EBV is responsible for some cases of post-transplant lymphoproliferative disorder, which represents the greatest risk of serious EBV disease in transplant recipients. HHV-6 and HHV-7 are two relatively newly discovered viruses and, at present, the body of information concerning these two agents is still fairly limited. KS is caused by HHV-8, which is the most recently discovered lymphotrophic HHV. Iatrogenic KS is seen in solid-organ transplant recipients, with a prevalence of 0.5-5% depending on the patient's country of origin. HPV is ubiquitous, and organ transplant recipients may never totally clear HPV infections, which are the most frequently recurring infections in renal transplant recipients. HPV infection in transplant recipients is important because of its link to the development of certain skin cancers, in particular, squamous cell carcinoma. Regular surveillance, sun avoidance, and patient education are important aspects of the management strategy.

Assessment of viral presence in semen and reproductive function of frozen-thawed spermatozoa from Pallas' cats (Otocolobus manul) infected with feline herpesvirus.

Although herpesviruses are known to contaminate the semen of several mammalian species, the occurrence of feline herpesvirus type 1 (FHV-1) in semen of infected cats has not been reported. Our objectives in this study were to investigate the presence of FHV-1 DNA in seminal fluid and frozen-thawed spermatozoa from FHV-1 infected Pallas' cats (Otocolobus manul) and assess the functionality of their frozen-thawed spermatozoa in vitro. Over a 3-yr period, semen (n = 33 ejaculates) was collected periodically via electroejaculation from four Pallas' cats chronically infected with FHV-1. Spermic ejaculates were frozen by pelleting on dry ice and stored in liquid nitrogen. After thawing, sperm motility and acrosome status were assessed over time during in vitro culture. For vitro fertilization (IVF), viable domestic cat (Felis silvestris catus) oocytes were inseminated with frozen-thawed Pallas' cat spermatozoa and evaluated for embryo cleavage. For FHV-1 polymerase chain reaction (PCR) analysis, DNA was extracted from seminal fluid, frozen-thawed spermatozoa, inseminated oocytes, heterologous IVF embryos, and conjunctival biopsies and analyzed for presence of a 322-base pair region of the FHV-1 thymidine kinase gene. Immediately post-thaw, sperm motility and percentage of intact acrosomes were decreased (P < 0.05) compared to fresh samples, and declined further (P < 0.05) during culture. However, all frozen-thawed IVF samples were capable of fertilizing domestic cat oocytes (overall, 46.1 +/- 6.0% cleavage). PCR analysis did not identify FHV-1 DNA in any reproductive sample despite the repeated detection of FHV-1 DNA in conjunctival biopsies. These results suggest that semen collected from Pallas' cats infected with FHV-1 does not contain cell-associated or non-cell-associated virus and that frozen-thawed spermatozoa exhibit adequate function for potential genetic rescue with minimal risk of FHV-1 transmission.

An inexpensive and rapid diagnostic method of Koi Herpesvirus (KHV) infection by loop-mediated isothermal amplification.

BACKGROUND: Koi Herpesvirus (KHV) affects both juvenile and adult common carp and koi, and is especially lethal to fry. The high mortalities caused by the disease have had a negative impact on the international koi trade. Different diagnostic techniques have been used to detect KHV, including: isolation of the virus in cell culture, electron microscopy, several PCR tests, ELISA and in situ hybridisation. All of these methods are time consuming, laborious and require specialised equipment. RESULTS: A rapid field diagnosis of KHV in common and koi carp was developed using loop-mediated isothermal amplification (LAMP). The LAMP reaction rapidly amplified nucleic acid with high specificity and efficiency under isothermal conditions using a simple water bath. Two methods of extracting DNA from host tissue were compared: extraction by boiling and by using a commercial extraction kit. A set of six primers--two inner primers, two outer primers and two loop primers--was designed from a KHV amplicon. The reaction conditions were optimised for detection of KHV in 60 min at 65 degrees C using Bst (Bacillus stearothermophilus) DNA polymerase. When visualised by gel electrophoresis, the products of the KHV LAMP assay appeared as a ladder pattern, with many bands of different sizes from 50 base-pairs (bp) up to the loading well. The KHV LAMP product could also be simply detected visually by adding SYBR Green I to the reaction tube and observing a colour change from orange to green. All samples positive for KHV by visual detection were confirmed positive by gel electrophoresis. The KHV LAMP had the same sensitivity as a standard PCR assay for the detection of KHV. CONCLUSION: This paper describes an accelerated LAMP assay for diagnosis of KHV. The entire procedure took only 90 minutes to produce a result: 15 minutes for DNA extraction; 60 min for the LAMP reaction; 2 min for visual detection using SYBR Green I. The test can be used under field conditions because the only equipment it requires is a water bath.

Yield and future issues of nucleic acid testing.

Despite the much lower actual yield than that estimated for hepatitis C virus (HCV) and human immunodeficiency virus (HIV) nucleic acid testing (NAT)-only positives in the USA and Germany, look-back procedures have revealed that no HCV transmission has occurred in Germany since the introduction of NAT. This indicates sufficient sensitivity of the pool-PCR approach. The slow ramp-up of hepatitis B virus (HBV) however, may require a different approach. It has been shown in Germany that the pooling of samples followed by virus enrichment results in a significant yield. Single donation testing for HBV would not increase the yield, because virus enrichment from mini-pool results in a similar sensitivity to that of single donation testing. Both strategies may be useful for extending future NAT to HBV screening. New candidate viruses for NAT are Parvo B19 and hepatitis A virus (HAV) because of their extreme resistance to inactivation procedures. Their low pathogenicity and epidemiologic characteristics, however, make them candidate viruses only for pooled source plasma. The main future issues of NAT will be related to the automation of pooling, extraction and amplification as a single homogeneous process. Depending on the throughput, automated single donation NAT as demonstrated by the 'Tigris' system may be an option, as far as all transfusion-relevant viruses will be included. In the near future high throughput systems will rely on pooled donor samples, most probably in conjunction with efficient enrichment procedures. For these systems, automation of the extraction and amplification process will be one of the first steps. These procedures will also limitthe costs of NAT and keep it available for use with future candidate viruses.

Application of in situ hybridization technique for quantitative assessment of ongoing symptomatic Epstein-Barr virus infection after living related liver transplantation.

For quantitative assessment of ongoing symptomatic Epstein-Barr virus (EBV) infection in pediatric recipients of liver transplantation, we determined the number of peripheral blood mononuclear cells (PBMC) infected by EBV by in situ hybridization (ISH) and related the results with clinical courses of those patients. Twenty-four patients had symptomatic EBV infection between February 1995 and March 1996. Blood samples were obtained from these 24 patients at the time of acute phase, from 13 of them during convalescence, and 37 pediatric patients before transplantation. ISH was performed on the PBMC and polymerase chain reaction (PCR) on DNA from whole blood. Oligonucleotide probes for ISH were chosen from coding sequences of EBV-encoded small nuclear RNA 1 (EBER1). Results of ISH were reported in a number of cells expressing EBER1/5 x 104 PBMC (#EBER1). Fever, diarrhea, upper respiratory symptoms, pleural effusion, ascites, lymphadenopathy, and lymphoproliferative disease (LPD) accompanied with EBV infection proven by serology, viral-specific stain or PCR were regarded as EBV related diseases (EBVD). All samples with positive #EBER1 were accompanied by positive EBV PCR. #EBERI was 68.2 +/- 144.9 (mean +/- SD) ranging from 0 to 621 in the acute phase, 0.20 +/- 0.41 ranging from 0 to 2 in the convalescence phase, 0.27 +/- 0.77 in 23 preoperative patients with positive serology, and 0 in all 14 preoperative patients with negative serology. The #EBER1 in ongoing EBVD was significantly greater than that of patients in convalescence or before transplantation. Patients with #EBERI greater than 10 had a significantly lower chance of convalescence and a higher mortality than patients with #EBER 1 less than 10. We conclude that #EBER1 could be a specific and quantitative marker of EBVD and might predict progression to LPD.

Serological and immunohistological assessment of Epstein-Barr virus infection in Sicilian patients with suspected nasopharyngeal carcinoma.

An evaluation of antibodies to Epstein-Barr virus (EBV) was carried out on 18 patients with suspected nasopharyngeal carcinoma (NPC). With one exception, 9 patients, with histologically confirmed NPC, had high levels of IgG and IgA antibodies to EBV-related antigens, VCA and EA. Out of 4 patients, without histologically confirmed NPC and an antibody pattern compatible with the disease, one developed NPC 22 months later. These data confirm the usefulness of serological markers as a diagnostic aid in NPC and indicate that the occurrence of this malignancy might be higher in Sicily than in low-risk zones.

Assessment of serologic markers for Epstein-Barr virus.

Antibody responses to early antigen (EA) and viral capsid antigen (VCA) were analyzed in 48 proven cases of Epstein-Barr Virus infection and in 48 age- and sex-matched healthy controls to establish optimal cutoff values for diagnosing EBV infection. Predictive values were determined for individual EA and VCA antibody titers and for EA to VCA antibody ratios and the optimal dilution cutoff values for positivity of EA (1:20), VCA (1:640), and EA to VCA (0:031) were selected. When evaluated on a subset of 10 VCA IgM positive cases and 35 negative controls, the three selected cutoff values identified as infections nine of 10, four of 10, and 10 of 10 of the cases and one of 35, none of 35, and one of 35 of the controls, respectively. When evaluated individually on 22 cases of suspected EBV infection who were heterophile antibody-negative and presented with symptoms compatible with EBV infection, an equal number of VCA IgM-positive and negative cases were identified as EBV infections. Overall, the cutoffs EA, VCA, and ratio identified 19 of 22 (86.4%), 14 of 22 (63.6%), and 18 of 22 (81.8%), respectively, and all cases could be identified using combinations of these values. Although these serologic values may be used with some accuracy, until more definitive markers are described a combination of heterophile responses, lymphocyte analysis, clinical symptoms, and serologic cutoff values should be used to assess the role of EBV in patient evaluation.

Primary virus isolation by a satellite laboratory.

A hospital-based satellite laboratory designed to isolate viral pathogens frequently found in renal transplant and hematology-oncology patients was compared with a system based on isolation by a state-wide reference laboratory. The hospital-based laboratory identified 12 viral isolates from the total 97 clinical specimens submitted. The hospital-based laboratory identified seven viral isolates from the first 50 clinical specimens, whereas the statewide reference laboratory identified six isolates from identical paired specimens. The average time from specimen collection to reporting results for the 97 specimens was 9.2 days for the hospital laboratory, as compared with 11.2 days for the first 50 duplicate specimens sent to the state laboratory. The cost per clinical specimen was $18.10, which compares favorably with costs for routine aerobic bacteriology. The use of a satellite laboratory system for primary viral isolation appears to provide rapid, accurate, and accessible viral diagnosis at an affordable cost.