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Background: La Crosse viral encephalitis (LACVE) is associated with residual epilepsy and neurocognitive deficits in survivors. This report summarizes 3 phases of clinical studies of children treated with intravenous (IV) ribavirin (RBV), each one exploring a different phase (I, IIA, IIB) of clinical trial development., Methods: In phase I, 7 children with life-threatening LACVE were treated with emergency use RBV using a moderate IV dose (8.33 mg/kg/dose q 8 hours day 1, 5 mg/kg/dose q 8 hours days 2-10). In phase IIA, 12 children with severe LACVE were enrolled: 8 treated with RBV (same dose as phase I) and 4 with placebo. In phase IIB an escalated dose was used (33 mg/kg dose 1, then 16 mg/kg/dose q 6 hours for 4 days, and 8 mg/kg/dose q 8 hours for 3 days)., Results: In a group of 15 children treated in phase I and phase IIA, RBV appeared safe at moderate dose, but based on steady-state RBV levels of 9.3 μM, estimated cerebrospinal fluid levels were less than 20% of the EC50 of RBV for LACVE. At the escalated dose used in phase IIB, adverse events occurred, likely related to RBV, and therefore the trial was discontinued. Nevertheless, valuable pharmacokinetic (PK) and safety data were obtained at moderate dose, with potential treatment implications for other indications., Conclusions: Although the results do not support the use of RBV for LACVE, this nevertheless is the largest study of antiviral treatment for LACVE to date and the largest pharmacokinetic analysis of IV RBV in children for any indication.

Objective: To describe factors associated with a rectal swab or stool sample positive for norovirus, rotavirus, or adenovirus., Design: Retrospective study., Setting: Charleston Area Medical Center, a regional academic medical center in Charleston, West Virginia., Methods: Rectal swab or stool samples were obtained from patients suspected of having viral gastroenteritis. These samples were sent to the Charleston Area Medical Center virology laboratory for testing in 2007. Viral antigen in rectal swab and stool samples is detected by use of commercially available immunoassay kits for each virus. Data were extracted from the virology laboratory database for the following 1-year time period: January 1, 2007, through December 31, 2007. When necessary, additional information was obtained from electronic administrative data on patients., Results: There were 2,867 rectal swab and stool samples available for viral testing. Of these samples, 1,261 (44%) were positive for a virus. Of these positive samples, 972 (77%) were positive for norovirus, 182 (14%) were positive for rotavirus, and 110 (9%) were positive for adenovirus. The patients in the youngest age group had the highest number of test results positive for all 3 viruses. When the test results for the youngest age group (0-9 years) were compared with those for all the other age groups combined (10-99 years), the proportion of positive cases was highest for the youngest age group (P<.001). There were significant seasonal trends for all 3 viruses. Multivariate analysis of norovirus showed that season, source, sex, and age were significant predictors of a positive test result. Multivariate analysis of rotavirus showed that season and source were significant predictors of a positive test result. Multivariate analysis of adenovirus showed that season and age were significant predictors of a positive test result., Conclusions: We conclude (1) that these 3 viruses are common causes of gastroenteritis in Charleston, West Virginia; (2) that infants and young children are more likely to test positive for these viruses than are older individuals; (3) that norovirus was the most common cause of gastroenteritis; and (4) that there are seasonal trends for all 3 viruses.

Data were collected on all patients in the Charleston, WV area tested for norovirus gastroenteritis during 2007. Of the 2687 rectal swab/stool samples, 60% were from individuals <20 years of age. Stool samples were more likely to be positive compared with rectal swab samples and if obtained from January to July and from patients <5 years of age.

Acute lower respiratory illnesses were prospectively investigated in a cohort of 1246 healthy infants who were enrolled at birth in the Tucson Children's Respiratory Study and followed through the first 3 years of life. Respiratory syncytial virus (RSV) infection was documented by culture, antigen detection or both in 276 episodes. In 21 (7.6%) of these 276, other viruses were simultaneously detected. Further serologic studies of 50 episodes in which RSV had been found increased the apparent viral codetection rate to 24%. When culture results for Chlamydia trachomatis and Mycoplasma pneumoniae were also considered, the rate of codetection was found to be 10.9% (30 of 276); this increased to 28% for the subgroup of episodes (14 of 50) that was further studied serologically. Illnesses associated with more than one agent were not significantly different from those involving RSV alone, with respect to month of onset, age at illness, illness type or duration of illness. We conclude that when RSV has been detected in previously healthy infants, routine searches for the concomitant presence of other viruses are usually not warranted.

Lower respiratory tract illnesses (LRIs) occurring during the first 3 years of life among children enrolled in the Tucson Children's Respiratory Study have been studied for evidence of viral, mycoplasmal and Chlamydia trachomatis infections. This report examines those from whom adequate acute and convalescent sera were available at the time of the LRI. Two groups were compared: those in whom culture and/or antigen detection yielded an etiologic agent (N = 110); and those who did not (culture negative, N = 124). Seroconversions (> or = 4-fold titer rise) to respiratory syncytial virus; influenza virus types A and B; parainfluenza virus types 1, 2 and 3; or adenovirus were found in only 0 to 5% of the culture negative group. No significant differences between groups with regard to frequencies of seroconversion to influenza type C, parainfluenza virus type 4, human coronaviruses 229E and OC43 or cytomegalovirus were detected, which suggests that these agents may not be frequent primary causes of LRIs among otherwise healthy children. Significant differences in seroconversions to Epstein-Barr virus were detected, suggesting that Epstein-Barr virus may contribute to LRI morbidity; however, its exact role remains to be defined.

Monoclonal antibody to measles virus was used successfully to identify measles virus antigen directly in clinical specimens, as well as in cell cultures. Pooled nasopharyngeal-throat swab specimens had a higher yield than throat swabs or urine samples for virus detection. Use of A549 cell cultures in the spin amplification vial assay proved to be highly efficient, allowing virus recognition within 1 to 2 days of inoculation. A combination of appropriately collected specimens, which includes a nasopharyngeal-throat swab, direct antigen detection with monoclonal antibody to measles in an indirect immunofluorescence system, and the spin amplification vial assay using A549 cells provides a sensitive and rapid system for isolation and/or identification of measles virus infections.

Pleomorphic, enveloped, virus-like particles were detected by electron microscopy in the stools of symptomatic infants during an outbreak of gastrointestinal illness in a neonatal intensive-care unit. To determine the incidence of virus-like particles in the stool and their relation to gastrointestinal symptoms, eight surveys of stools for the particles were conducted over 40 weeks. The incidence of virus-like particles in the stool decreased from 69% to less than 10% over the study period. Most infants surveyed were premature; overall, 32 (36%) of 88 neonates were positive for virus-like particles. Statistically significant associations were found between virus-like particles in the stool and gastrointestinal symptoms within one week of each survey. These symptoms included water-loss stools, blood in the stool, gastric retention, bilious gastric aspirates, and abdominal distention. Several infants with virus-like particles whose mothers had gastrointestinal or "flu-like" symptoms before delivery were identified in the community (not part of the survey study).

During an 8-yr period, 862 stool specimens from patients with gastroenteritis were examined by electron microscopy after negative staining with 2% phosphotungstic acid (pH 6.5). Forty-one percent of the specimens submitted over an 8-yr period were determined to be positive for virus or viruslike particles belonging to one or more of seven morphologically distinct viral groups. Coronavirus-like particles (CVLPs) were present in 69.8% of the positive stool specimens. Membranous profiles containing "complement-type" holes (10 nm in diameter) were identified in some preparations containing CVLPs. The second most prevalent viral agent found in stool specimens was the rotavirus (17% of all positive stools). The incidence of other viruses identified in the survey were as follows: adenovirus 4.5%, picorna/parvovirus agents 2.9%, Norwalk-like agent 2.9%, astrovirus 1.9%, and calicivirus 0.5%. Unclassified small round viruses (approximately 25-30 nm in diameter) represented 0.5%. It was also determined that there was a seasonal distribution in excretion of all viruses except for CVLPs. A greater number of viruses were identified in the cooler, drier months of the year.

Laboratory strains of group B coxsackievirus serotypes 1 to 6 were inoculated intraperitoneally into newborn mice of differing genetic backgrouns. Of the four genetic strains investigated, C3H mice appeared to be resistant to all six serotypes, whereas BALB/c mice were most susceptible. Swiss mice and a random-bred Swiss strain (COH) were intermediate in susceptibility. The findings underscore the fact that clinical isolation attempts and experimental studies involving group B coxsackieviruses must take into account both the virus strain used and the genetic background of the host. For clinical isolation of these viruses, the BALB/c mouse may be the most suitable of th strains tested.

Coronaviruslike particles (CVLPs) were visualized by direct electron microscopy (EM) of diarrheal stools in 49 of 126 infants and children between 1 month and 12 years of age during a three-year observation period. The clinical and epidemiologic features of these patients were analyzed and compared with patients with diarrhea who were shedding rotaviruses, or whose stools were negative for enteric viruses by EM. Seasonal and age distributions for CVLP shedding were similar to those for rotaviruses (in most cases less than 1 year of age; peak months were September through January), as were the symptoms and median durations of illness. Prospective studies of three subsequent patients suggest that the duration of shedding in acute illness is five to at least 25 days. Multiple attempts to cultivate the CVLPs were unsuccessful. In addition, partial purification of CVLPs from stool specimens was performed, and immunologic analysis by immunoelectron microscopy and radial immunodiffusion showed no antigenic relatedness to prototype human (OC43 and 229E) or animal (bovine and canine) coronaviruses. These findings suggest that CVLPs may be an important cause of acute gastrointestinal illness in infancy, and may represent a virus antigenically unrelated to known human and animal coronaviruses.

A fluorescein isothiocyanate-conjugated pool of monoclonal antibodies (MoAb) to respiratory syncytial virus (RSV) was prospectively evaluated for its utility as a direct, 1-hr test for the diagnosis of RSV infection. Direct nasopharyngeal swab smears collected from 109 infants and children with acute respiratory illnesses were studied and compared with results obtained by indirect immunofluorescence using bovine polyclonal anti-RSV antibody on eluted cells derived from pooled nasopharyngeal and throat swab specimens (a 2.5-3 hr procedure), and culture. The MoAb-direct smear method was at least 86%-89% sensitive and 95%-100% specific compared with either of the other procedures. Additional prospective evaluations, as well as retrospective studies on a selected bank of slides stored from the preceding year, established that this MoAb could also be used with confidence in testing where direct smears are not employed.

Immunofluorescence staining methods for respiratory syncytial virus antigen detection were compared. Of 50 specimens originally positive for respiratory syncytial virus by direct immunofluorescence and culture, 49 were positive by repeat direct immunofluorescence and 32 were positive by indirect immunofluorescence. Additional results obtained on specimens originally negative for respiratory syncytial virus by direct immunofluorescence, culture, or both indicate that direct immunofluorescence staining for respiratory syncytial virus antigen was more sensitive than was indirect immunofluorescence.

Quantitation of T and B lymphocytes in infants and children is an important test in the diagnosis of a suspected immuno-deficiency. Previous studies indicated that the absolute and relative numbers of lymphocyte subpopulations vary with age, but these data in the pediatric age group are incomplete and often contradictory. We reviewed the literature and investigated the relationship between age and lymphocyte subpopulations in healthy infants and children using common methods and recent methodologic improvements. We found that absolute numbers of T and B cells followed the same trend as the total lymphocyte count, which was elevated at birth, increased in the first six months, and then gradually decreased to adult levels at approximately 13 years of age. Compared with adult values, the percentage of B cells also was higher at birth and continued to increase for six months, followed by a gradual decrease to adult levels by late childhood or early adolescence. The percentage of T cells gradually increased to adult levels by the same age range.

An illness lasting for two years, with recurrent fever, rash, abdominal pain, and arthralgia, developed in a four year old boy. He was found to have a combined Epstein-Barr virus and cytomegalovirus (CMV) infection. His symptoms, CMV in his urine, and an absent in vitro lymphocyte response to CMV antigen persisted for two years. After treatment with orally administered bovine transfer factor clinical symptoms and viruria disappeared and specific immunity to CMV developed. Evaluation of this treatment in chronic virus infections is warranted.

Serum varicella-zoster (VZ) antibody titers were determined by a simple, indirect immunofluorescent antibody technique in 50 children with leukemia or non-Hodgkin's lymphoma. Sixteen children had initial antibody titers that were not detectable at a serum dilution of 1:8 and were considered to be susceptible to varicella. Thirty-one patients had initial serum VZ antibody titers of 1:16 or greater, while three had levels of 1:8. The serum antibody titer was 1:8 or greater in 16 children with a history of varicella. Seven episodes of localized herpes zoster were observed in children whose VZ titer was 1:8 or greater prior to onset. Two nonfatal infections with primary varicella developed in children with leukemia in remission whose initial titers were less than 1:8 and were associated with a convalescent rise in VZ antibody to levels greater than 1:16. Three susceptible children received zoster immune plasma or zoster immune globulin for varicella exposure, causing a transient rise in serum VZ titer. The assessment of serum VZ titer by this immunofluorescent antibody technique combined with the prior varicella history is useful in defining the population of children with leukemia and non-Hodgkin's lymphoma who are susceptible to primary varicella.

Rapid immunofluorescence (FA) methods for the detection of common respiratory viruses were compared with culture results over a 3-year period to assess the relative efficiency of FA in a clinical laboratory setting. For respiratory syncytial virus, efficiencies were high (sensitivity, 90 to 95%; specificity, 92 to 95%). The sensitivity of FA for detection of parainfluenza virus type 1, parainfluenza virus type 3, influenza A virus, and adenoviruses ranged from 28 to 63%, but specificities for these viruses were uniformly 98 to 100%. The observations form a basis for consideration of selective reduction of routine culture procedures for specimens with initial positive rapid FA results; however, the possibility of dual viral infection in some situations must also be considered.

We studied three patients with influenza virus-associated fulminant myocarditis; one was infected by type B and the others by type A influenza virus. In one patient, dissemination of type A (H1N1) virus to the myocardium was demonstrated, and viremia complicated the clinical course despite the use of oral amantadine HCl and ribavirin aerosol. All patients were treated with iv ribavirin, two initially and the third after viremia was detected during hyperacute rejection of a cardiac transplant. No significant adverse effects could be directly attributed to therapy, and viral shedding abruptly terminated coincident with its use; however, both patients treated shortly after onset of myocarditis died. The third required support by an artificial heart, and died 8 mo later. Immunotyping of myocardial tissues in two cases revealed an initial predominance of T helper cells. Serial endomyocardial biopsies available from one of these demonstrated a subsequent marked decrease in the T helper cell population as inflammation and necrosis subsided during and following therapy.

This study examines the relationship between age and mitogen-induced lymphocyte blastogenesis. Lymphocytes were stimulated with PHA; Concanavalin A (Con A), and PWM in 156 normal, healthy subjects ranging in age from birth to 75 years. The findings indicate a gradual but significant decrease in PHA- and Con A-induced blastogenesis with increasing age. The decrease in Con A and PHA induced in vitro lymphocyte function begins in early childhood and young adulthood, respectively, and continues throughout the age range studied. In addition, there appears to be a decreased range of lymphocyte functional capacity among the 50-75-year-old subjects. The clinical and laboratory implications of these observations are discussed.

This study examines the relationship between percent recovery of lymphocytes and T- and B-cell typing results. Lymphocytes were recovered from heparinized whole blood by density gradient centrifugation. T- and B-cells were enumerated by spontaneous sheep rosetting and direct fluorescent antibody staining for surface immunoglobulin, respectively. The findings indicate that at low recovery levels ( less than 80%) there is a significant increase in variability of lymphocyte typing results and a significant increase in IgG-bearing B-lymphocytes. Possible explanations for these findings are discussed. These results suggest that lymphocyte typing results should be interpreted in relation to recovery level and that recovery levels of greater than or equal to 80% are desirable.

We describe two patients with serologic evidence of active Epstein-Barr virus infection who presented with unusual neurologic manifestations and minimal systemic findings of infectious mononucleosis. One girl developed an acute hemiplegic migraine syndrome followed by acute psychosis, and the other patient had severe, generalized chorea. The wide spectrum of presenting central nervous system findings associated with Epstein-Barr virus infections underscores the need to suspect this agent in a variety of acute neurologic syndromes.

Forty-four patients, including 26 adults and 18 children under 15 years of age, were referred for evaluation of recurrent or persistent illnesses, with symptoms including pharyngitis, lymphadenopathy, fever, headaches, arthralgia, fatigue, depression, dyslogia, and myalgia. Thirty-nine patients were positive for Epstein-Barr virus antibody with antibody levels compatible with active infection for at least 1 year. Antiviral capsid antigen and anti-early antigen titers of patients were significantly greater (p less than 0.001) than age-group-matched controls. The frequency, number, duration, and patterns of symptoms, as well as patient sex, were compared by age in study patients seropositive and seronegative for Epstein-Barr virus. Illness patterns were not associated with changes in specific antibody titers or clinical findings. Lymphocyte phenotype and function analyses were done in 11 of the 39 patients positive for Epstein-Barr virus antibody; no consistent differences from normal were found. Only 1 of 32 patients had circulating interferon, in contrast to 7 of 7 patients with acute infectious mononucleosis. There were many adverse consequences of the illness. Epstein-Barr virus infection may not be self-limiting, and the virus may be associated with clinically recognizable illness other than infectious mononucleosis in children as well as in adults.

A high-passage laboratory strain of coxsackievirus B1 produced a unique myositis that predominantly and profoundly affected hip extensors and, to a lesser extent, hindquarter knee flexors when inoculated into a strain of random-bred newborn mice of Swiss origin (COH mice). The effect was not observed in BALB/c or C3H mice similarly inoculated. In addition to the differences in susceptibility of the mouse strains, it was found that six different low-passage "field" isolates of coxsackie-virus B1 isolated from infected patients varied considerably in their capability to induce these lesions. Thus, selective myositis which is muscle group-specific can be induced in a mouse model with coxsackievirus B1, and both genetic factors of the host and characteristics of the virus strain play a significant role in the pathogenesis of the myositis.

A 55-year-old man had a myelopathic syndrome and serologic evidence of Epstein-Barr virus infection. The patient's age and the neurologic and serologic findings suggested reactivation of a latent virus with direct invasion of the CNS. Epstein-Barr virus may exist in a latent state within the nervous system, and should be considered in myelopathic syndromes, even in older patients.

Nine patients with an initial onset of symptoms of acute arthritis within the preceding four weeks were enrolled in a prospective serological study with clinical follow-up for six months to two years. Four adults with chronic rheumatoid arthritis and ten healthy adults were similarly studied. Serial titers measured included antibodies to Epstein-Barr virus (EBV) antigens, group B coxsackieviruses, rubella virus, cytomegalovirus, and herpes simplex virus. Serological evidence of active EBV infection was found in four of the patients with acute arthritis, none of the patients with chronic arthritis, and one of the ten healthy adults. There was no similar correlation between acute disease and presence of antibodies to the other viruses tested. We suggest that EBV may cause acute rheumatic illnesses more commonly than is currently appreciated but is probably not involved in the etiology of typical chronic rheumatoid arthritis.

Hemadsorption of primary monkey kidney cell cultures was commenced at 1 day after inoculation to evaluate how rapidly influenza A virus, influenza B virus, and parainfluenza virus types 1, 2, and 3 could be detected by this method from respiratory specimens. Overall, 38% of all isolates could be detected within 24 h of inoculation, and 69% could be detected within 48 h. All influenza A viruses and all but one influenza B virus were detected by day 3.

Indirect immunofluorescence and microneutralization methods for the detection of antibodies to poliovirus serotypes 1, 2, and 3 were compared. Of the 41 sera tested for poliovirus type 1 antibody, 40% were in complete agreement, 55% differed by one dilution, and 5% differed by two dilutions. For poliovirus type 2, 37 sera were tested; 56% completely agreed, and 44% differed by only one dilution. For poliovirus type 3, complete agreement occurred in 59% of 33 sera, while 41% differed by one dilution. No false-negative results were obtained. These findings suggest that indirect immunofluorescence for poliovirus types 1, 2, and 3 is as sensitive as the microneutralization method and could represent a less cumbersome alternative.