Detection Of Latent Infection Research Articles

Peeling the Onion: Towards a Better Understanding of Botrytis Diseases of Onion.

Onion is cultivated worldwide for its bulbs, but production is threatened by pathogens and pests. Three distinct diseases of onion are caused by species that belong to the fungal genus Botrytis. Leaf blight is a well-known foliar disease caused by B. squamosa that can cause serious yield losses. Neck rot is a postharvest disease that manifests in bulbs after storage and is associated with three species: B. aclada, B. allii, and B. byssoidea. The symptomless infection of onion plants in the field makes it difficult to predict the incidence of neck rot in storage, although progress on the detection of latent infection has been made. In onion cultivation for seed production, blighting of the inflorescence is caused by all four onion-specific Botrytis species plus the broad host range pathogen B. cinerea. Flower blight can reduce seed yield and contaminate seed. In this review, the long history of Botrytis diseases of onion is discussed, as well as recent and future approaches to acquire a better understanding of the biology and ecology of Botrytis spp. pathogenic on onion. New fundamental insights in the genetic, biochemical, and physiological aspects of Botrytis-onion interactions are essential to improve the breeding of Botrytis-resistant onion cultivars.[Formula: see text] Copyright © 2021 The Author(s). This is an open access article distributed under the CC BY-NC-ND 4.0 International license.

Molecular Detection of Ralstonia solanacearum to Facilitate Breeding for Resistance to Bacterial Wilt in Potato.

Potato bacterial wilt is caused by the devastating bacterial pathogen Ralstonia solanacearum. Quantitative resistance to this disease has been and is currently introgressed from a number of wild relatives into cultivated varieties through laborious breeding programs. Here, we present two methods that we have developed to facilitate the screening for resistance to bacterial wilt in potato. The first one uses R. solanacearum reporter strains constitutively expressing the luxCDABE operon or the green fluorescent protein (gfp) to follow pathogen colonization in potato germplasm. Luminescent strains are used for nondestructive live imaging, while fluorescent ones enable precise pathogen visualization inside the plant tissues through confocal microscopy. The second method is a BIO-multiplex-PCR assay that is useful for sensitive and specific detection of viable R. solanacearum (IIB-1) cells in latently infected potato plants. This BIO-multiplex-PCR assay can specifically detect IIB-1 sequevar strains as well as strains belonging to all four R. solanacearum phylotypes and is sensitive enough to detect without DNA extraction ten bacterial cells per mL in complex samples.The described methods allow the detection of latent infections in roots and stems of asymptomatic plants and were shown to be efficient tools to assist potato breeding programs.

A quick and efficient hydroponic potato infection method for evaluating potato resistance and Ralstonia solanacearum virulence

BackgroundPotato, the third most important crop worldwide, plays a critical role in human food security. Brown rot, one of the most destructive potato diseases caused by Ralstonia solanacearum, results in huge economic losses every year. A quick, stable, low cost and high throughout method is required to meet the demands of identification of germplasm resistance to bacterial wilt in potato breeding programs.ResultsHere we present a novel R. solanacearum hydroponic infection assay on potato plants grown in vitro. Through testing wilt symptom appearance and bacterial colonization in aerial part of plants, we found that the optimum conditions for in vitro potato infection were using an OD600 0.01 bacterial solution suspended with tap water for infection, broken potato roots and an open container. Infection using R. solanacearum strains with differential degree of aggressivity demonstrated that this infection system is equally efficient as soil-drench inoculation for assessment of R. solanacearum virulence on potato. A small-scale assessment of 32 potato germplasms identified three varieties highly resistant to the pathogen, which indicates this infection system is a useful method for high-throughout screening of potato germplasm for resistance. Furthermore, we demonstrate the utility of a strain carrying luminescence to easily quantify bacterial colonization and the detection of latent infections in hydroponic conditions, which can be efficiently used in potato breeding programs.ConclusionsWe have established a quick and efficient in vitro potato infection system, which may facilitate breeding for new potato cultivars with high resistance to R. solanacearum.

Detection of Latent Peronospora effusa Infections in Spinach.

Downy mildew disease of spinach, caused by Peronospora effusa, is managed in conventional fields by a combination of host resistance and scheduled fungicide applications. Fungicides are currently applied to prevent downy mildew epidemics regardless of the infection status of spinach crops. A more streamlined approach would be to develop methods to target either latent infections for fungicide application in conventional production systems or to hasten harvest in organic production. In this study, conventional polymerase chain reaction (PCR) was applied to detect P. effusa DNA in symptomless spinach leaves in three spatially and temporally separated field plots, each containing four 2-m beds, 35 m in length. Spinach leaves were sampled weekly at 3-m intervals at 48 locations throughout each plot. Initial samples were asymptomatic and yet PCR enabled detection of P. effusa DNA extracted from sampled spinach leaves. Detection of latent downy mildew infection in spinach leaves was confirmed by PCR as early as 7 days prior to symptom development. The limit of pathogen DNA detection in spinach leaves was calculated at 10 pg using the conventional PCR approach. Quantitative PCR with TaqMan methodology revealed the presence of inhibitors from spinach leaf DNA extracts and affected amplification efficiencies, but not when diluted, enabling detection of P. effusa DNA at a concentration of <0.1 pg. In conclusion, detection of latent infections may enable management decisions for earlier-than-normal harvest of infected, symptomless organic crops, and for timing fungicide applications on symptomless plants in conventional production.

Detection of latent infections caused by Colletotrichum sp. in olive fruit.

To set up a practical method to detect latent infections of Colletotrichum sp., the causal agent of olive anthracnose, on olives before the onset of disease symptoms. Freezing, sodium hydroxide (NaOH), ethanol and ethylene treatments were evaluated to detect latent infections on inoculated and naturally infected olive fruit by Colletotrichum sp. as non-hazardous alternatives to paraquat. Treatments were conducted using fruit of cultivars Arbequina and Hojiblanca. The disease incidence and T50 were calculated. Dipping in NaOH 0·05% solution and the paraquat method were the most effective treatments on both inoculated and naturally infected fruit, although the value of T50 was lower for the NaOH method than for the paraquat method in one of the experiments. Subsequently, the dipping time in NaOH 0·05% was evaluated. Longer dipping times in NaOH 0·05% were better than shorter ones in cultivar Arbequina, with 72h being the most effective in cultivar Hojiblanca. NaOH solution is a practical method to detect latent infections of Colletotrichum sp. on immature olive fruit. This study is relevant because we set up a viable, non-hazardous alternative to paraquat to detect latent infections of Colletotrichum sp. using NaOH. The use of NaOH is a simple and eco-friendly tool that allows the determination of the level of latent infections by Colletotrichum in olives. Therefore, our method will be useful in decision-making processes for disease management before the appearance of the first visible symptoms.

Development and validation of loop mediated isothermal amplification based detection assay for Puccinia striiformis f. sp. tritici causing stripe rust of wheat

Wheat stripe rust caused by Puccinia striiformis f. sp. tritici is one of the most important diseases of wheat worldwide. Detection of latent infection in leaves is critical for estimating the initial inoculum potential for causing epidemics. A loop-mediated isothermal amplification (LAMP) assay was developed and validated for the detection of P. striiformis f. sp. tritici DNA in urediniospores and wheat seedlings with latent infections. Reaction temperature, optimal Mg2+ concentration and ratio of outer/inner primers for reaction efficiency were optimized. The sensitivity of detection was 1 picogram of genomic DNA and marker could detect latent infection in wheat leaves as early as 48 h post inoculation. This colorimetric detection assay was indicated by color change from purple to violet color in positive reaction by adding 200 μM hydroxyl napthol blue solution (HNB) to the reaction mixture. LAMP products were further analyzed by visualizing EtBr stained reaction mixture on transilluminator that showed full brightness in positive reaction which was absent in negative reactions. Further confirmation of LAMP products through agarose gel electrophoresis showed ladder like banding pattern only in positive reactions. The developed method is efficient, rapid and sensitive for detection of Puccinia striiformis tritici causing stripe rust of wheat. No cross reaction with other related pathogens was observed. The developed technique is very useful in quantifying inoculum load during offseason which would help in development of prediction model and also be useful in development of diagnostic kit.

Interspecific Potato Breeding Lines Display Differential Colonization Patterns and Induced Defense Responses after Ralstonia solanacearum Infection.

Potato (Solanum tuberosum L.) is one of the main hosts of Ralstonia solanacearum, the causative agent of bacterial wilt. This plant pathogen bacteria produce asymptomatic latent infections that promote its global spread, hindering disease control. A potato breeding program is conducted in Uruguay based on the introgression of resistance from the wild native species S. commersonii Dun. Currently, several backcrosses were generated exploiting the high genetic variability of this wild species resulting in advanced interspecific breeding lines with different levels of bacterial wilt resistance. The overall aim of this work was to characterize the interaction of the improved potato germplasm with R. solanacearum. Potato clones with different responses to R. solanacearum were selected, and colonization, dissemination and multiplication patterns after infection were evaluated. A R. solanacearum strain belonging to the phylotype IIB-sequevar 1, with high aggressiveness on potato was genetically modified to constitutively generate fluorescence and luminescence from either the green fluorescence protein gene or lux operon. These reporter strains were used to allow a direct and precise visualization of fluorescent and luminescent cells in plant tissues by confocal microscopy and luminometry. Based on wilting scoring and detection of latent infections, the selected clones were classified as susceptible or tolerant, while no immune-like resistance response was identified. Typical wilting symptoms in susceptible plants were correlated with high concentrations of bacteria in roots and along the stems. Tolerant clones showed a colonization pattern restricted to roots and a limited number of xylem vessels only in the stem base. Results indicate that resistance in potato is achieved through restriction of bacterial invasion and multiplication inside plant tissues, particularly in stems. Tolerant plants were also characterized by induction of anatomical and biochemical changes after R. solanacearum infection, including hyperplasic activity of conductor tissue, tylose production, callose and lignin deposition, and accumulation of reactive oxygen species. This study highlights the potential of the identified tolerant interspecific potato clones as valuable genetic resources for potato-breeding programs and leads to a better understanding of resistance against R. solanacearum in potato.

Species-specific PCR-based assays for identification and detection of Botryosphaeriaceae species causing stem blight on blueberry in China

Botryosphaeriaceae species are important causal agents of blueberry stem blight worldwide. Blueberry stem blight has become an important disease, potentially affecting the quality and production of blueberries in China. It is difficult and time-consuming to identify at the species level using morphological methods. The aim of this study was to develop polymerase chain reaction (PCR) assays for the diagnosis and early detection of latent infections of blueberry stems by Botryosphaeria spp. Species-specific primers, based on the ribosomal DNA internal transcribed spacer region and β-tubulin gene, were designed and selected for use in PCR assays. Three primer pairs, Lt347-F/R for Lasiodiplodia theobromae, Np304-F/R for Neofusicoccum parvum and FaF/Bt2b for Botryosphaeria dothidea, successfully amplified specific PCR fragments of different sizes on pure cultures or from blueberry stems inoculated and naturally infected blueberry plants with three pathogens, respectively. These primers did not amplify any PCR fragments from other blueberry stem disease-associated pathogens, such as Phomopsis spp. and Pestalotiopsis spp. This PCR protocol could detect as low as 100 pg to 1 ng of purified fungal DNA. This PCR-based protocol could be used for the diagnosis and detection of these pathogens from pure cultures or from infected blueberry plants.

Excretory Secretory Proteins Released during Growth of Mycobacterium tuberculosis (H37Ra), With Diagnostic Potential in Pulmonary and Extra Pulmonary Tuberculosis

TB immunodiagnostics based on antibody and antigen detection for early detection and monitoring tubercular infections at low cost and flexible to adapt to field laboratories are a boon to developing countries. In this context we have explored in-house developed penicillinase based ELISA assays for the detection of antigen, antibody as well as immune-complexed antigen using various excretory secretory antigenic proteins and specific antibodies. Excretory secretory protein antigens such as ES-31, ES-41, ES-43, ES-6, ES-20, ES-100 and EST-6 were studied extensively and found to be useful in various pulmonary and extra pulmonary cases of tuberculosis infection. ES-31 has shown its diagnostic potential in chronic PTB cases, ES-43 in relapse cases and ES-41 in bone and joint TB. Elevated level of ES-20 antigen was observed in patients with weak immune response in TB lymphadenitis. Detection of ES-100 antigen and antibody by penicilinase ELISA was observed to be useful in detection of TB meningitis. ES-6 antigen was shown to be useful in detection of latent infection. These proteins with antigenic properties are utilized for production of specific antibodies against them and observed to be useful in immune diagnostics for detection of specific circulating and immune complexed antigens. Further user friendly peroxidase immunoassay has been standardised and is being routinely used for suspected TB cases attending 1000 bedded Kasturba Hospital attached to medical institute on physician’s request.

Controle do inóculo inicial para redução dos danos pela podridão: 'olho-de-boi' em macieiras

O objetivo deste trabalho foi avaliar o impacto da redução do inóculo no controle da podridão-'olho-de-boi' (POB) causada por Cryptosporiopsis sp. em maçãs e minimização de perdas. Os experimentos foram realizados em Vacaria-RS, nos ciclos 2006/07 e 2007/08. Na fase de dormência de macieiras Pink Lady®, foram aplicados os seguintes tratamentos erradicantes (dose p.i.): calda sulfocálcica (3,0%); oxicloreto de cobre (0,5%); hidróxido de cobre (0,3%); óxido cuproso (0,3%) e testemunha sem pulverização. A redução do inóculo foi avaliada em solução oriunda da lavagem de ramos, gemas floríferas e frutos, semeados em meio semisseletivo. Na colheita, a incidência da POB foi avaliada em quatro categorias de frutos: caídos sob a área de projeção da copa; nas plantas; infecções latentes, e após três meses de armazenamento em câmara frigorífica. Houve relação linear positiva entre os níveis de inóculo de Cryptosporiopsis sp. em gemas e os danos por POB (R² = 0,948; P = 0,001). O uso da calda sulfocálcica possibilitou a maior redução de inóculo nas gemas, com redução em 50% dos danos totais ocasionados por POB na safra de 2006/07. Nos frutos, o inóculo apresentou um incremento linear ao longo do ciclo, sendo relativamente maior em 2007/08, onde os tratamentos erradicantes possibilitaram controle superior a 40% nos danos com benefício econômico pela redução do inóculo.

Development and implementation of cost-effective strategies to manage brown rot of peach trees in Imathia, Greece

Development of cost-effective strategies to manage brown rot caused by Monilinia laxa of peach implies a better understanding of the susceptibility of different cultivars and pre-harvest contamination. This study investigated the susceptibility of 24 peach and nectarine cultivars to shoot blight caused by Monilinia spp. and found various levels of susceptibility, with the nectarine cultivar Tasty Free scored as the most susceptible. Studies on the the existence and detection of latent infections by Monilinia spp. in three peach (‘A37’, ‘Andross’, and ‘E-45’) and three nectarine (‘Venus’, ‘Fantasia’, and ‘Tasty Free’) cultivars were also conducted. The results showed that latent infections were detected only in nectarine cultivars when fruit were collected on 23 May and 22 June. In contrast, nectarine fruit collected on 7 June and all peach cultivars tested had no detectable latent infection. This study also indicated that the fungicide thiophanate methyl applied at the pit hardening stage reduced significantly the percentage of latent infection and subsequently preharvest fruit rots. Finally, a disease forecast model to predict blossom blight, caused by M. laxa, was evaluated in the Prefecture of Imathia, Greece. Trees, sprayed according to the model predictions, showed a statistically lower percentage of blighted shoots than those of unsprayed trees.

Cocoa swollen shoot virus in Ghana: A review of diagnostic procedures

A quick and more reliable diagnostic method has for a long time been identified as one input that will greatly enhance the control of the cocoa swollen shoot disease in Ghana. Many diagnostic procedures have been developed for detecting the virus that causes the disease; yet, the detection of latent infections is still unpredictable. This paper reviews work done in diagnosing swollen shoot disease to develop a quicker, more reliable, and cost-effective method that could be applicable in programmes for controlling cocoa swollen shoot disease. Enzyme-linked immunosorbent assay (ELISA), virobacterial agglutination tests (VBA), and other biochemical analyses have been discussed as diagnostic tools that could be developed to effectively replace the use of visible symptom expressions as the only method for field detection of the disease.

Detection of latent infections in mango fruit with herbicides

The aim of this study was to study the effect of herbicides on the breakdown of mango fruit to identify the fungi recovered from the herbicides-treated fruit and to determine the potential use of herbicides in a rapid quantitative measure of the pathogenic component of the fruit storability. Quick detection of symptomless latent anthracnose of mango was used as nine herbicides. It may be seen that all the herbicides tested were found induce the manifestation of latency earlier than the control. It is interesting to note that paraquat (3000 ppm) was able to manifest the disease (64.33%) within 3–5 days versus 7–10 days in control followed by Basalin (36.66%) and Alachlor (34.00%). Butacholar and Pretalacholar were not effective even over 6–10 days in inducing the manifestation of latent infection. For fruits treated with paraquat, one can detect the disease much earlier and decide whether they are fit enough for long-term storage, export or quick disposal in the local market.

Multiplex PCR and quality control of Epinotia aporema granulovirus production

A specific multiplex PCR was developed for the rapid and highly sensitive quality control of the viral DNA during Epinotia aporema granulovirus (EpapGV) production. At the beginning of this work only 2.3% of the EpapGV genomic sequence was known. In order to increase the availability of specific information, the terminal sequences of the inserts of several selected clones of EpapGV genomic libraries were determined. These data comprised 8.4% of the total DNA sequence and corresponded to regions distributed throughout the genome. Based on the small fraction of known sequence available a set of 32 primers was designed, using information theory to set the basis for this study. Each pair of designed primers was initially tested in individual PCRs to assess the correct size of the expected product and the sensitivity of the amplification. The specificity was verified in multiplex PCRs, using alternatively 1-3 sets of selected 5-6 primer pairs and EpapGV DNA preparations from different sources and degrees of purity. The results indicate that the multiplex PCR could be used for quality control in the bioinsecticide production, as well as in other applications such as the detection of latent infections in E. aporema colonies, and studies related to virus distribution, vertical transmission, host range, or persistence in the field.

Survey of tuberculosis incidents in hospital healthcare workers, England and Wales, 2005

Tuberculosis (TB) incidence has increased in England and Wales since 1987, with much of the increase in migrants from high TB incidence countries. The National Health Service increasingly depends on healthcare workers (HCWs) trained abroad, often from areas of high TB incidence. A retrospective survey of TB incidents in hospital-based HCWs was carried out to describe their frequency, distribution and characteristics. Reports of HCWs with TB in hospitals were identified among routine surveillance of TB incidents. Additional data on these and reports of further incidents fitting the study definition were obtained by contacting local and regional Health Protection Units. At least 105 incidents of TB in hospital-based HCWs occurred in England and Wales in 2005. Most involved HCWs from high incidence countries, and most cases had pre-employment occupational health screening. We found no evidence of onward transmission within hospitals. Pre-employment screening for active disease may not be enough to prevent the occurrence of these incidents. A high index of suspicion among HCWs with TB symptoms is needed. Detection of latent infection with interferon gamma release assays, and the use of preventive treatment, should be evaluated.

Antemortem detection of latent infection with neuropathogenic strains of equine herpesvirus-1 in horses

Antemortem detection of latent infection with neuropathogenic strains of equine herpesvirus-1 in horses

DNA based detection of blackleg and soft rot disease causing Erwinia strains in seed potatoes

Erwinia carotovora subsp. atroseptica (Eca), Erwinia carotovora subsp. carotovora (Ecc) and Erwinia chrysanthemi (Ech) are the different sub species of Erwinia that cause the diseases commonly known as blackleg, aerial stem rot and soft rot on potato. Blackleg and aerial stem rot affect vines during the growing season, whereas soft rot affects tubers in the field and during transit and storage. The three species can cause soft rot under cool and moist conditions. E. carotovora subsp. atroseptica is the major cause of blackleg, a blackening of the stem base of potato plants, which originates from the mother tuber (Pérombelon and Kelman, 1987). Erwinia carotovora subsp. carotovora mainly causes aerial stem rot (aerial blackleg), but under high temperatures it has been reported to cause blackleg like symptoms. E. chrysanthemi also induces blackleg-like symptoms. Until recently E. chrysanthemi had been mainly confined to warmer climates of Europe, Australia and the tropics. To date the species has been known to occur in cool temperate regions including Finland. In Finland E. chrysanthemi the strain has been reported for the first time in 2004 and it appears to spread fast in certain parts of country (Degefu, unpublished, Pirhonen, personal communication). Although infested crop residues and rotting tubers are among the important sources of inoculum, latent infections in seed tuber provide the major source of infection in potato production (Hannukkala and Segertedt, 2004). At the seed potato laboratory MTT, Ruukki we are carrying out research and services on PCR (DNA) based detection of latent infection of blackleg and soft rot Erwinia strains. The enrichment of the bacteria in semi selective liquid medium prior to PCR ( BIO-PCR) is an important initial step for the success in PCR detection extremely low number of the target bacteria from tubers. The different strains appear to differ in their ability to compete with other saprophytes and reach the target detection limit of bacterial population during the enrichment culture of the potato peel extract. Results of prior PCR enrichment of the bacteria, detection limits of the different strains and preliminary data, from the analysis of some seed lots from the high grade area of north Ostrobothnia and Åland regions, on the occurrence of the strains and the new trends of introduction and spread of E. chrysanthemi in Finland are presented and discussed. Evaluation of the current status of Erwinia diagnostics and areas of future research are highlighted

Detection and Quantification of Botrytis cinerea by ELISA in Pear Stems During Cold Storage.

Botrytis cinerea was detected and quantified in pear stems from six orchards in the Pacific Northwest, and changes in fungal biomass in the stems after 6 and 8 months of cold storage in regular (air) atmosphere were studied. The fungus was detected by plating stem halves on selective medium and by enzyme-linked immunosorbent assay (ELISA) using the Botrytis-specific monoclonal antibody BC-12.CA4. Both methods demonstrated that the incidence of B. cinerea increased from 6 to 8 months, but ELISA was more sensitive than standard isolation. Quantitative ELISAs on stems indicated that over 200 μg of B. cinerea biomass per gram of stem tissue was present in the stems of visibly rotted fruits, but usually less than 35 μg/g was present in stems from fruits without visible gray mold. Aureobasidium pullulans, Penicillium spp., Alternaria spp., and Cladosporium spp. were commonly isolated from stem tissue. A. pullulans was present in 86% of the stems from which B. cinerea was detected. Use of the monoclonal antibody BC-12.CA4 could help in determining the infection path of B. cinerea in pear stems and detection of latent infections, enabling the timing and method of control of stem end rot to be optimized.

Cell Block Preparation of a Burkitt’s Lymphoma Cell Line as a Positive Control for in Situ Hybridization for Epstein-Barr Virus

To examine the feasibility of the use of a cell block preparation of Epstein-Barr virus (EBV)-infected Burkitt's lymphoma cell line (EB-3) as a positive control for in situ hybridization (ISH) for EBV-encoded RNA (EBER). EB-3 cells were processed into cell block and cytocentrifuge preparations. ISH for EBER was performed, and the results were compared with a commercially available EBV positive control slide (cytocentrifuge). The cost of preparing the cell block and cytocentrifuge sample was only a fraction of the price of commercial control slides. ISH for EBER was strongly stained in all preparations with similar intensity of staining but with a much higher number of positive cells in the cell block. Although the cellular details in the cell block were considerably inferior to those on the cytocentrifuge and commercial control slide, with distortion of nuclei and smudging of chromatin, the interpretation of the ISH results was unaffected. The cell block was stable at room temperature when examined after one year. The cell block preparation of an EBV-infected Burkitt's lymphoma cell line serves as a reliable, stable and a relatively inexpensive control, with good preservation of cellular details of cellular RNA for ISH study for EBER in the detection of latent infection with EBV.

Detection of latent infection by Epstein-Barr virus in peripheral blood cells of healthy individuals and in non-neoplastic tonsillar tissue from patients by reverse transcription-polymerase chain reaction.

A highly sensitive and specific reverse transcription-polymerase chain reaction (RT-PCR) assay was designed for detection of Epstein-Barr virus (EBV)-related sequences in nucleic acid extracted by the conventional phenol method. Using this EBV-infected malignant lymphoma cell line, Raji cells, a comparative study of this assay was carried out with EBER1 (EBV-encoded small RNA1) primer and conventional DNA-PCR with BamHI-W and EBER1 primers, respectively. The results revealed that this assay has sensitivity about 10(5)-fold higher than the conventional DNA-PCR method. The presence of EBER1 DNA and RNA was also investigated in 23 healthy individuals and 22 right and left tonsils of 11 healthy individuals. These results indicated that this assay is both sensitive and specific. Thus, EBV infection could be diagnosed easily determined and EBER1 was shown to be transcribed in peripheral blood cells and tonsils with quantitatively different grades. This assay can be used to diagnose EBV infection in clinical samples.