Rabbit Kidney Cell Cultures Research Articles

Comparative characterization of Leporipoxvirus members’ reproduction in continuous cell culture

Examination of the virus-cell interactions is of both scientific and practical importance. Our study was aimed at comparative characterization of rabbit myxoma virus and Shope fibroma virus biological properties that manifested during the virus reproduction in RK-13/2-03 clonal continuous rabbit kidney cell culture. It was demonstrated that the viruses varied in infection development and cytopathic effect duration in RK-13/2-03 cell culture. Apparent lesions in cell monolayers infected by myxoma virus and fibroma virus at similar multiplicity of infection and cultivation temperature were observed on day 2 and day 3 of cultivation, respec- tively, as well as maximum cell lesions with evident degeneration were observed on day 3 and day 6 of cultivation, respectively. Myxoma virus was accumulated at titre of 6.25–6.50 lg TCID 50 /0,2 cm 3 , and Shope fibroma virusа was accumulated at titre of 5.50−5.75 lg TCID 50 /0.2 cm 3 . Shope fibroma virus demonstrated such infectivity during three passages and myxoma virus demonstrated such infectivity during twenty passages. Prepared cultures were identified as myxoma virus and Shope fibroma virus with molecular genetic analysis. Tests of the viruses for their antigenic relatedness showed that antibodies against myxoma virus were able to neutralize Shope fibroma virus also. NT titres of antibodies against both viruses were similar (1:8). RK-13/2-03 cell culture was found to be highly permissive to Shope fibroma virus that had been isolated from the diseased rabbit and not been an attenuated variant.

Significant upregulation of U1 and U4 spliceosomal snRNAs by ATP nanoliposomes explains acceleration of wound healing, due to increased pre-mRNA processing to functional mRNA

Delayed wound healing is one of the hallmarks of diabetic complications and certain autoimmune inflammatory diseases. Extensive wound healing studies in rabbits have indicated that the delivery of ATP encapsulated in unilamellar nanoliposomes causes rapid cell proliferation and fast tracks the wound healing process. In the current study, we explored the possible molecular mechanism underlying this response by comparing gene expression in cultured rabbit kidney cells treated with either ATP nanoliposomes (containing 1 mg Mg-ATP/ml formulation) or control nanoliposomes (containing 1 mg/ml unmetabolisable gamma-thio-ATP/ml formulation). High-quality total RNA was isolated 24 h from the cells and subjected to RNA seq technology, which revealed significant overexpression of specific noncoding RNAs. The U1 spliceosomal RNA, U1 snRNA, was upregulated more than 250-fold following treatment with ATP nanoliposomes. This multifunctional U1 spliceosomal RNA may function in transcription by speeding up the critical splicing step, thereby facilitating faster processing of pre-mRNA to translatable mRNA.

ВИВЧЕННЯ ПРОТИГЕРПЕТИЧНОЇ АКТИВНОСТІ ТАБЛЕТОК АЛЬТАБОР

<p><strong>STUDY OF ANTIHERPETIC ACTIVITY OF TABLETS ALTABOR </strong></p><p><strong>Т.V. Кrutskikh<sup>1</sup>, N.V. Nesterova<sup>2</sup>, S.D Zagorodnya<sup>2</sup> </strong></p><p><sup>1</sup>National Pharmaceutical University, Kharkiv</p><p> <sup>2</sup>Institute of Microbiology and Virology NAS of Ukraine, Kyiv, Ukraine</p><p><strong>Summary:</strong> studied antiviral activity of tablets Altabor. Determined that the drug has antiherpetic activity as therapeutic and prophylactic use of schemes.</p><p><strong>Key words: </strong>altabor, antiherpetic activity, herpes, tablets.<strong></strong></p><p><strong>Introduction.</strong> Common classification of acute and recurrent herpes human diseases currently still pending. This is due to the fact that while patients with this disease are not treated to a specialist, and the doctors of different specialties - depending on the localization of acute and recurrent processes, resulting in clinical practice herpes virus disease diagnosis is "topical" in nature, etiology the disease is ignored and software etiopathogenetic treatment is performed. Today herpes infection is attracting more attention as the medical and non-medical circles of society. This is due to the fact that cold sores, known since ancient times as small and insignificant spread of disease, recently greatly expanded its importance in human pathology. Increased interest in herpes infection is also due to several other reasons and advance all that has now spread to different parts of the world, strikes to 95% of the population, and the virus is capable of causing damage almost all the organs and systems of the human body and cause various forms of infection - acute, latent and chronic recurrent. Clinically herpes occurs as diverse, complex and often severe disease with lesions of many organs and tissues, which can be considered as a general systemic disease organism.</p><p>Modern medicine has therapies that allow to eliminate the herpes simplex virus from the human body. Until recently, treatment was generally symptomatic herpes. Prescribed remedies that dried, to anesthetize, cooled and prevented infection lesions other microorganisms. Tried to apply and antivirals (ointment oksolin, tebrofen), but the effectiveness of most antiviral agents left much to be desired. Today treat herpes infections is complex and characterized by low efficiency. Therefore, therapies aimed at preventing the development or restoration of disorders that cause activation of the herpes virus in the body. At this time there are two main directions in the treatment of herpes, it is, first, the use of etiopathogenetic antiviral therapy, the main place in which to play acyclic nucleoside - acyclovir (Zovirax, acyclovir-acre), valaciclovir (Valtrex) and famciclovir (famvir) and Secondly, a comprehensive treatment that includes immunotherapy (specific and nonspecific) in combination with antiviral therapy.</p><p>Number of drugs for the treatment of herpetic lesions is limited because of new original products is appropriate and timely. One of these drugs is original domestic product - Altabor tablets that are based on Altabor substance that has antiherpetic and erferonogenie action.</p><p><strong>Methods.</strong> Antiherpetic activity of Altabor tablets studied in models in vitro and in vivo.</p><p><strong>Results and discussion</strong>. To study antiherpetic tablets altabor activity in vitro using rabbit kidney cell culture infected with the herpes virus type 1. The results demonstrated that both in medical and in prevention of various concentrations of tablets altabor (from 100 to 6.2 mcg/ml) was observed expressed antiherpetic activity. If preventive action tablets altabor minimum active concentration was 6.2 mg/ml, for therapeutic action - 3.1 mg/ml.</p><p>Antiherpetic activity of altabor tablets studied in vivo against the herpes simplex virus 1 model herpes virus meningoencephalitis in white outbred mice. As the reference product used "Viroleks» KRKA (Slovenia). Based on these data concluded that the tablets have expressed altabor preventive effect at a dose of 0.5 mg/kg per os, better protect mice from herpes meningoencephalitis than drug Viroleks. Also, based on certain performance index (IE=60) can assert pronounced therapeutic activity of altabor tablets, which are much more efficient than Viroleks.</p><strong>Conclusions.</strong> Results of the studies on the antiviral activity of tablets Altabor found that the drug shows pronounced antiherpetic activity as a therapeutic and prophylactic regimens application. If preventive action tablets altabor minimum active concentration was 6.2 mg/ml, the therapeutic action - 3.1 mg/ml. At a dose of 0.5 mg/kg per os altabor more effectively protects mice from herpes meningoencephalitis than drug Viroleks, 2 times better at prevention and 1.5 times in the application of medical schemes.

Relevance of the Interaction between Alphaherpesvirus UL3.5 and UL48 Proteins for Virion Maturation and Neuroinvasion

The UL3.5 and UL48 genes, which are conserved in most alphaherpesvirus genomes, are important for maturation of pseudorabies virus (PrV) particles in the cytoplasm of infected cells (W. Fuchs, B. G. Klupp, H. J. Rziha, and T. C. Mettenleiter, J. Virol. 70:3517-3527, 1996; W. Fuchs, H. Granzow, B. G. Klupp, M. Kopp and T. C. Mettenleiter, J. Virol. 76:6729-6742, 2002). In bovine herpesvirus 1 (BoHV-1), the homologous gene products pUL3.5 and pUL48 have been demonstrated to interact physically (N. Lam and G. Letchworth, J. Virol. 74:2876-2884, 2000). Moreover, BoHV-1 pUL3.5 partially complemented a pUL3.5 defect in PrV (W. Fuchs, H. Granzow, and T. C. Mettenleiter, J. Virol. 71:8886-8892, 1997). By using coimmunoprecipitation and yeast two-hybrid studies, we observed a similar interaction between pUL3.5 and pUL48 of PrV, as well as a heterologous interaction between the PrV and BoHV-1 gene products. The relevant domain could be confined to the first 43 amino acids of PrV pUL3.5. Unlike its BoHV-1 homologue, PrV pUL3.5 is processed by proteolytic cleavage, and only an abundant 14-kDa fragment consisting of amino acids 1 to >or=116 could be detected by peptide mass fingerprint analysis of purified wild-type PrV particles, which also contain the pUL48 tegument component. To determine the biological relevance of the protein-protein interaction, pUL3.5-, pUL48-, and double-negative PrV mutants were analyzed in parallel. All deletion mutants were replication competent but exhibited significantly reduced plaque sizes and virus titers in cultured rabbit kidney cells compared to wild-type and rescued viruses, which correlated with a delayed neuroinvasion in intranasally infected mice. Remarkably, the defects of the double-negative mutant were similar to those of pUL48-negative virus. Electron microscopy of cells infected with either deletion mutant revealed the retention of naked nucleocapsids in the cytoplasm and the absence of mature virus particles. In summary, our studies for the first time demonstrate the relevance of the pUL3.5-pUL48 interaction for secondary envelopment of an alphaherpesvirus, give a molecular basis for the observed trans-complementation between the PrV and BHV-1 pUL3.5 homologs, yield conclusive evidence for the incorporation of a proteolytically processed pUL3.5 into PrV virions, and demonstrate the importance of both proteins for neuroinvasion and neurovirulence of PrV.

Isolation of Equine Herpesvirus-5 from Blood Mononuclear Cells of a Gelding

Horses are commonly infected by herpesviruses, but isolation of equine herpesvirus-5 (EHV-5) has only infrequently been reported. We describe the isolation and characterization of a strain of EHV-5 from the blood mononuclear cells of a healthy adult horse in California. The virus was initially identified by EHV-5 specific polymerase chain reaction (PCR), and it caused lytic infection of cultured rabbit kidney cells only after repeated serial passage. Virions with characteristic herpesvirus morphology were readily demonstrated in cell culture lysate by transmission electron microscopy. A portion of the glycoprotein B gene of this strain of EHV-5 had 99% identity to the published EHV-5 sequence, and it was clearly distinguishable from other EHV (1-4) by virus-specific PCR assays. Prevalence of EHV-5 infection in a group of young racehorses was estimated at 64% using the EHV-5 specific PCR on nasopharyngeal secretions.

Identification and Characterization of Two Subpopulations of Encephalitozoon intestinalis

Microsporidia are obligate intracellular protozoa that have been shown to be pathogenic to most living creatures. The development of in vitro cell culture propagation methods has provided researchers with large numbers of spores and facilitated the study of these organisms. Here, we describe heterogeneity within cell culture-propagated Encephalitozoon intestinalis suspensions. Flow cytometer histograms depicting the log side scatter and forward-angle light scatter of spores from nine suspensions produced over 12 months consistently showed two populations differing in size. The suspensions were composed primarily of the smaller-spore subpopulation (76.4% +/- 5.1%). The presence of two subpopulations was confirmed by microscopic examination and image analysis (P < 0.001). Small subpopulation spores were noninfectious in rabbit kidney (RK13) cell culture infectivity assays, while the large spores were infectious when inocula included > or = 25 spores. The small spores stained brilliantly with fluorescein isothiocyanate-conjugated monoclonal antibody against Encephalitozoon genus spore wall antigen, while the large spores stained poorly. There was no difference in staining intensities using commercial (MicroSporFA) and experimental polyclonal antibodies. Vital-dye (DAPI [4',6'-diamidino-2-phenylindole], propidium iodide, or SYTOX Green) staining showed the spores of the small subpopulation to be permeable to all vital dyes tested, while spores of the large subpopulation were not permeable in the absence of ethanol pretreatment. PCR using primers directed to the 16S rRNA or beta-tubulin genes and subsequent sequence analysis confirmed both subpopulations as E. intestinalis. Our data suggest that existing cell culture propagation methods produce two types of spores differing in infectivity, and the presence of these noninfective spores in purified spore suspensions should be considered when designing disinfection and drug treatment studies.

The Effects of Elevated Temperatures and Various Time‐Temperature Combinations on the Development of Brachiola (Nosema) algerae N. Comb. in Mammalian Cell Culture

Nosema algerae Vávra and Undeen 1970, a microsporidian known to cause infection in mosquitoes, develops in mammalian cell cultures at 24-35 degrees C and in the tails and footpads of athymic mice. More recently it has been reported to grow at 38 degrees C in human cell culture. The present study is a two-part temperature/development examination. The first part examines the development of N. algerae in rabbit kidney cell culture at 29 degrees C, which permits the formation of functional spores within 72 h, and compares the effect of elevated temperatures (36.0, 36.5, 37 degrees C) on parasite development. At these elevated temperatures, N. algerae infects but undergoes only one or two proliferative divisions, with no evidence of sporogony by 72 h post-inoculation. During this time, however, the host cells continue to divide resulting in fewer infected cells over time and giving the appearance of a diminished parasitemia. Additionally, at 37 degrees C some organisms degenerate/hibernate by 72 h while others remain viable/active. It is not until 96 h that the parasites appear in large clusters of proliferative stages in the few host cells that are infected. By 120 h post-inoculation, proliferative cells, sporoblasts, and early spores are observed. These results suggest that elevated temperatures impede proliferation rates and the onset of sporogony. The second part of this study evaluates developmental changes in N. algerae when incubation temperatures and times are varied during parasite growth, resulting in abnormal parasite morphology. These abnormalities were not present when parasites were grown at constant temperature (29-37 degrees C). This report demonstrates that N. algerae can successfully develop at high temperatures (37 degrees C), justifying its taxonomic relocation to the genus Brachiola.

A spore counting method and cell culture model for chlorine disinfection studies of Encephalitozoon syn. Septata intestinalis.

The microsporidia have recently been recognized as a group of pathogens that have potential for waterborne transmission; however, little is known about the effects of routine disinfection on microsporidian spore viability. In this study, in vitro growth of Encephalitozoon syn. Septata intestinalis, a microsporidium found in the human gut, was used as a model to assess the effect of chlorine on the infectivity and viability of microsporidian spores. Spore inoculum concentrations were determined by using spectrophotometric measurements (percent transmittance at 625 nm) and by traditional hemacytometer counting. To determine quantitative dose-response data for spore infectivity, we optimized a rabbit kidney cell culture system in 24-well plates, which facilitated calculation of a 50% tissue culture infective dose (TCID(50)) and a minimal infective dose (MID) for E. intestinalis. The TCID(50) is a quantitative measure of infectivity and growth and is the number of organisms that must be present to infect 50% of the cell culture wells tested. The MID is as a measure of a system's permissiveness to infection and a measure of spore infectivity. A standardized MID and a standardized TCID(50) have not been reported previously for any microsporidian species. Both types of doses are reported in this paper, and the values were used to evaluate the effects of chlorine disinfection on the in vitro growth of microsporidia. Spores were treated with chlorine at concentrations of 0, 1, 2, 5, and 10 mg/liter. The exposure times ranged from 0 to 80 min at 25 degrees C and pH 7. MID data for E. intestinalis were compared before and after chlorine disinfection. A 3-log reduction (99.9% inhibition) in the E. intestinalis MID was observed at a chlorine concentration of 2 mg/liter after a minimum exposure time of 16 min. The log(10) reduction results based on percent transmittance-derived spore counts were equivalent to the results based on hemacytometer-derived spore counts. Our data suggest that chlorine treatment may be an effective water treatment for E. intestinalis and that spectrophotometric methods may be substituted for labor-intensive hemacytometer methods when spores are counted in laboratory-based chlorine disinfection studies.

Induction of metallothionein synthesis by cadmium and zinc in cultured rabbit kidney cells (RK-13).

The effects of increasing concentrations of Zn(II) and Cd(II) on the expression of the four isometallothioneins (isoMTs), namely MT-1a, MT-2a, MT-2d and MT-2e, in rabbit kidney cells (RK-13) and the development of cellular tolerance to these metal ions were studied. The results showed that, whereas in parental cells MT concentration was low and composed nearly exclusively of MT-2a and MT-1a, all four isoMTs increased massively in abundance when the cells were exposed to toxic concentrations of Zn(II) or Cd(II), the relative increase being largest in the two minor isoforms MT-2d and MT-2e. While the response of the four isoMTs to the challenge by Zn(II) or Cd(II) was qualitatively comparable, there were differences in sensitivity and delay time, Cd(II) being the more efficient inducer and much faster in eliciting the onset of isoMT synthesis. An even larger production of isoMTs resulted when RK-13 cells were cultured in the presence of a series of metal concentrations yielding sub-lines of increased metal tolerance. In this instance too, there were marked differences in the response to Cd(II) and Zn(II). Thus, in cells of sub-lines selected for tolerance to moderate concentrations of Cd(II) the kinetic analysis of isoMT accretion gave indications of a saturable induction process while no such evidence was forthcoming for Zn(II). In sub-line cells selected for tolerance to the highest concentrations of Cd(II) or Zn(II) isoMT formation was increased by another order of magnitude, reaching for some isoforms a 100- to 1000-fold augmentation over the amounts measured in cells of the unexposed parental cells. A potentiation of this magnitude goes beyond the range of ordinary regulation of gene expression. It is to be viewed instead as an enlargement of the capacity of isoMT synthesis acquired by a variety of mechanisms in the surviving cells.

Resolution and Quantification of Four Metallothionein Isoforms from Rabbit Kidney Cells

Reversed-phase high-performance liquid chromatography coupled with an on-line integrating chromatographic data system was used to separate and quantify the four major isometallothioneins MT-1a, MT-2a, MT2d, and MT-2e present in metallothionein samples from cultured rabbit kidney cells as prepared by gel-filtration and/or ion-exchange chromatography. The standard curves for each of these isoforms showed closely comparable linear correlations between the amount of protein applied to the column and their integrated absorbance peak area at 220 nm. The lower limit of quantification is 30 pmol, sufficient to assess basal isometallothionein concentrations and to follow their variation upon metal exposure.

Studies on serum renotropic activity after uninephrectomy in rabbits.

The present study was undertaken to critically examine the hypothesis that circulating renotropin is the primary stimulus for compensatory renal growth. Blood was collected before and after either left nephrectomy or sham operation from a total of 34 rabbits and the serum renotropic activity assessed by the uptake of 3H-thymidine in primary cultures of rabbit kidney cells. Sera obtained after uninephrectomy had significantly more renotropic activity than sera collected before surgery. In some cases, sera obtained after sham operation had more renotropic activity than those collected before, but the mean response was not increased after sham operation. The difference between sera collected before and after uninephrectomy was not transferrable to primary cultures of rabbit liver cells. These data confirm a modest but significant influence of uninephrectomy on the renotropic activity of blood in rabbits.

Comparison of standard culture methods, a shell vial assay, and a DNA probe for the detection of herpes simplex virus

A nonradioactive, biotinylated herpes simplex virus (HSV) DNA probe, a shell vial (rabbit kidney cell) culture assay enhanced by a direct fluorescent (HSV monoclonal)-antibody stain at 16 to 20 h postinoculation, and conventional tube cultures with confirmation via HSV-specific (polyclonal antibody) immunoperoxidase assay were compared for 199 specimens. The predictive values of the positive results were 54.5% for the probe, 95.9% for the shell vial assay, and 100% for the conventional culture methods, while the predictive values of the negative tests were 68.1, 84.0, and 98.4%, respectively. We conclude that the DNA probe (sensitivity, 24.5%; specificity, 88.3%) and the shell vial assay (sensitivity, 66.2%; specificity, 98.4%) cannot be substituted for conventional tube culture techniques (sensitivity, 97.1%; specificity, 100%) in the routine identification of HSV in our laboratory.

Serum renotropic factor stimulates prostaglandin synthesis in primary cultures of rabbit kidney cells

Compensatory growth of the kidney occurs in response to a partial reduction in renal mass. This compensatory renal growth may be regulated by a circulating renotropic factor. Prostaglandin synthesis has been shown to be increased in kidneys undergoing compensatory renal growth in vivo. In the present study we observed that the addition of rabbit sera obtained after uninephrectomy enhanced DNA synthesis in primary cultures of rabbit kidney cells compared to sera obtained prenephrectomy. The stimulated kidney cells produced more prostaglandin E 2 than control cells. Furthermore, the addition of prostaglandin E 2 to rabbit kidney cells in the presence of control sera also stimulated DNA synthesis. These results provide further evidence that prostaglandins may participate in the biological events which regulate renal growth in response to a circulating renotropic factor.

Induction of neutralizing antibodies against bovine leukosis virus in rabbits by vaccination with recombinant vaccinia virus expressing bovine leukosis virus envelope glycoprotein

SUMMARY Three kinds of recombinant vaccinia virus (rvv)—mO-ha/ati, LO1-ha/ati and mO-ha/7.5kD—expressing bovine leukosis virus (blv) envelope glycoprotein (gp60) were constructed. The blv envelope gene of rvv mO-ha/ati and LO1-ha/ati or of rvv mO-ha/7.5kD was expressed under control of the promoter of A-type inclusion body (ati) protein gene of cow-pox virus or vaccinia virus 7.5-kD protein gene, respectively. The vaccinia virus strain, LC16mO, was used as vector for rvv mO-ha/ati and mO-ha/7.5kD, and strain LO-1 was used for rvv LO1-ha/ati. Strains LC16mO and LO-1 are attenuated vaccine virus strains originating from the Lister original vaccinia virus. All 3 kinds of constructed rvv expressed gp60 in cultured rabbit kidney cells after infection; mO-ha/ati expressed more antigen than did mO-ha/7.5kD. Rabbits vaccinated with rvv produced considerable antibody capable of inihibiting syncytium formation, as well as antibody with virion-binding ability. The rvv that used ati promoter induced higher antibody titer than did the rvv that used 7.5-kD promoter. Results indicate that blv gp60 is responsible for induction of neutralizing antibodies that suppress in vitro formation of syncytia among blv-infected cells. Applicability of rvv, especially those using ati promoter, was evaluated in a vaccine against bovine leukosis.

Experimental Transmission and Electron Microscopic Demonstration of the Virus of Haemorrhagic Disease of Rabbits in Czechoslovakia

Intramuscular administration of the filtrate of organ suspensions, prepared from a dead rabbit, killed 62.9% of inoculated rabbits within 1 to 5 days, while 93.3% died after intranasal administration of the same inoculum. The virus survived freeze-drying and was resistant to treatment with 0.4% formaldehyde when incubated at 37 degrees C for 1 hour and 4 degrees C for the subsequent 12 hours, but lost its infectivity when the treatment was prolonged to 3 hours at 37 degrees C and 3 days at room temperature. Its infectivity was also inhibited by reconvalescent serum. The virus could not be detected after 3 passages in primary rabbit kidney cell cultures. Electron microscopy of negatively stained preparations demonstrated icosahedral virus particles with a diameter of 29 to 33 nm without an envelope. Accurate morphological classification has not yet been completed. Incubation with a reconvalescent serum, diluted 1:20 or 1:40, resulted in the formation of immune complexes, detectable by electron microscopy.

A Cost-effective Method for Recovery of Herpes Simplex Virus

Specimens submitted for recovery of herpes simplex virus (HSV) were each inoculated to two Cultureset (CS) cultures and a single primary rabbit kidney (RK) cell culture. The first CS was stained after 24 to 48 hours at 35°C. The second CS was similarly processed at three to four days if the first CS was negative. RK cultures were incubated at 35°C for six days with daily inspection for cytopathic effects (CPEs). All CS cultures and only those RK cultures with CPEs were stained with a direct immunoperoxidase procedure. RK cells were stained within the tubes. HSV was detected in 95 (18%) of the 529 specimens processed. Of the positive specimens, 77 (81%) were found with CS within two days, and 13 (14%) more were detected at three or four days; 90 (95%) were found with RK, 46 (48%) within two days. Inexperienced technologists can be easily trained to detect CPEs from HSV in RK cultures. A combination of one CS and one RK cell culture per specimen would provide a rapid, comprehensive, and cost-effective alternative to the routine use of two CS cultures per specimen.

Detection of Herpes Simplex Virus With Primary Rabbit Kidney and an HSV Antigen ELISA

A total of 411 specimens submitted for herpes simplex virus (HSV) was used to evaluate primary rabbit kidney cell cultures and an HSV antigen (culture-amplified) enzyme-linked immunosorbent assay (ELISA). Primary rabbit kidney cell cultures were compared against routinely used cell cultures of primary rhesus monkey kidney (PRMK), primary human embryonic kidney (PHEK), diploid human lung fibroblast (WI-38). and whole human embryo (Flow-5000). The HSV antigen ELISA was evaluated against primary rabbit kidney cultures, which compared favorably with routinely used cell cultures. It detected 91.7% of HSV positives by day 4 and the rest within seven days. Additionally, primary rabbit kidney cell cultures detected two more HSV positives not previously detected by the other cell cultures. The HSV antigen ELISA presented a 90.7% sensitivity and 97.9% specificity in detecting HSV-positive cultures. Combined use of primary rabbit kidney cell cultures and the HSV antigen ELISA has eliminated our use of expensive cell cultures and blind passages for HSV cultures; has decreased reporting time of negative reports from 14 to 4 days, and has been cost-effective in reducing the time and effort needed for HSV identification.

Comparison of commercial sources of primary rabbit kidney and MRC-5 cell cultures for herpes simplex virus isolation

Two commercial sources of MRC-5 and primary rabbit kidney cell cultures were compared for sensitivity of herpes simplex virus isolation and cytopathic effect detection time. Forty-six of 78 previously positive and 50 of 234 fresh clinical specimens were positive and could be evaluated. Herpes simplex virus recovery from frozen specimens positive in at least one cell culture ranged from 85% in MRC-5 cells (M.A. Bioproducts) to 89% in primary rabbit kidney cells (M.A. Bioproducts). Herpes simplex virus recovery from fresh clinical specimens ranged from 94% in primary rabbit kidney cells (Flow Laboratories), MRC-5 cells (Flow Laboratories), and MRC-5 (M.A. Bioproducts) cells to 100% in primary rabbit kidney (M.A. Bioproducts) cells. Among the 96 positive specimens evaluated, no significant difference was found between commercially prepared MRC-5 and primary rabbit kidney cell cultures in either sensitivity or time to first detectable cytopathic effect. Cells from M.A. Bioproducts and Flow Laboratories were equally sensitive. Herpes simplex virus detection was dependent upon inoculating at least two cell culture tubes rather than if the cells were MRC-5 cells or primary rabbit kidney cells.

Synthesis and antiviral activity of acyclic derivatives of 5-ethyl-2'-deoxyuridine.

AbstractFour acyclic pyrimidine derivatives of 5‐ethyl‐2′‐deoxyuridine (1) (EDU) have been synthesized and tested for their activities against vesicular stomatitis virus, vaccinia virus and various strains of herpes simplex virus type 1 (HSV‐1) and herpes simplex virus type 2 (HSV‐2) in primary rabbit kidney (PRK) cell cultures. The acyclic nucleoside analogues of EDU exhibited no activity against any of the viruses tested.

Prodrugs of 5-ethyl-2'-deoxyuridine, II. Syntheses and antiviral activities of 5'- and 3'-ester derivatives.

AbstractWith the aim of obtaining derivatives of the well established antiherpes compound 5‐ethyl‐2′‐deoxyuridine (1) (Aedurid©, EtUdR, EDU), which are more lipophilic and therapeutically superior, 5′‐ and 3′‐ester derivatives of 1 were synthesized. Tested in primary rabbit kidney cell cultures against various strains of herpes simplex type 1 (HSV‐1) and type 2 (HSV‐2), all EtUdR esters, with the exception of compounds 8 and 13, proved almost as active as EtUdR itself, suggesting that they were readily hydrolized.