[Risk factors associated with bacterial growth in derivative systems from cerebrospinal liquid in pediatric patients]. / Factores de riesgo asociados a crecimiento bacteriano en sistemas derivativos de líquido cefalorraquídeo en pacientes pediátricos.

OBJECTIVE: To determine risk factors associated with bacterial growth in systems derived from cerebrospinal fluid in pediatric patients. METHODS: Case and controls study from January to December 2012, in patients aged <16 years who were carriers of hydrocephalus and who required placement or replacement of derivative system. Cases were considered as children with cultures with bacterial growth and controls with negative bacterial growth. Inferential statistics with Chi-squared and Mann-Whitney U tests. Association of risk with odds ratio. RESULTS: We reviewed 746 registries, cases n=99 (13%) and controls n=647 (87%). Masculine gender 58 (57%) vs. feminine gender 297 (46%) (p=0.530). Age of cases: median, five months and controls, one year (p=0.02). Median weight, 7 vs. 10 kg (p=0.634). Surgical interventions: median n=2 (range, 1-8) vs. n=1 (range, 1-7). Infection rate, 13.2%. Main etiology ductal stenosis, n=29 (29%) vs. n=50 (23%) (p=0.530). Non-communicating, n=50 (51%) vs. 396 (61%) (p=0.456). Predominant microorganisms: enterobacteria, pseudomonas, and enterococcus. Non-use of iodized dressing OR=2.6 (range, 1.8-4.3), use of connector OR=6.8 (range, 1.9-24.0), System replacement OR=2.0 (range, 1.3-3.1), assistant without surgical facemask OR=9.7 (range, 2.3-42.0). CONCLUSIONS: Being a breastfeeding infant, of low weight, non-application of iodized dressing, use of connector, previous derivation, and lack of adherence to aseptic technique were all factors associated with ependymitis.

Fulminant ependymitis following intraventricular rupture of brain abscess.

A 48-year-old man with a history of a penetrating brain injury was referred with a presumptive diagnosis of bacterial meningitis. Examination revealed a brain abscess in addition to meningitis. Blood and cerebrospinal fluid (CSF) cultures were negative for bacteria, and empirical IV antibiotic therapy with vancomycin (VCM) and meropenem was initiated. Despite initial improvement, however, his condition rapidly deteriorated into coma following intraventricular rupture of the abscess and hydrocephalus. Thereafter, an emergency ventriculostomy was performed and the abscess was evacuated. Bacterial cultures of the pus were negative. To manage the hydrocephalus, 150-200 ml of CSF were drained daily. Intraventricular administration of VCM (20 mg q.d.) was added to the IV antibiotic therapeutic regimen after surgery. Although the primary abscess rapidly decreased in size, ependymitis developed in the fourth ventricle. This new lesion, which resulted from CSF dissemination from the primary abscess, was refractory to treatment, and eventually disappeared after the intraventricular VCM dosage was increased from 20 to 30 mg and continued for 30 days. A possible reason for the development of fulminant ependymitis and why it was refractory to treatment despite the shrinkage of the primary lesion may be that physiological CSF flow from the lateral to the fourth ventricle was lost due to CSF drainage, and the stagnant CSF flow coupled with an insufficient VCM level in the fourth ventricle facilitated the rapid growth of pathogens. Although intraventricular antibiotic administration is efficacious for treating ruptured brain abscesses, it may be associated with the unexpected development of secondary lesions.

The behaviour of both Listeria monocytogenes and rat ciliated ependymal cells is altered during their co-culture.

BACKGROUND: Ciliated ependymal cells line the cerebral ventricles and aqueducts separating the infected CSF from the brain parenchyma in meningitis. PRINCIPAL FINDINGS: Investigation of the interaction of Listeria monocytogenes with cultured rat brain ependymal cells showed that certain strains reduced the beat frequency of the cilia but all the strains studied significantly reduced the ciliary beat amplitude (the linear distance travelled by the tip of each cilium per beat cycle). CONCLUSION: The presence of the ependyma caused aggregation of some listeria strains and in some cases extracellular material also was seen in association with bacterial aggregates. These observations were dependent on the expression of genes required for invasion, intracellular survival and listerial cell to cell spread that are regulated by the transcriptional activator, positive regulatory factor A (PrfA).

Stenosis of the central canal of the spinal cord following inoculation of suckling hamsters with reovirus type I.

The central canal of the human spinal cord is partially or completely occluded in the vast majority of individuals by the early years of adult life. The authors describe an experimental lesion following virus-induced ependymitis that bears a striking resemblance to the condition in man. Suckling hamsters were inoculated with 0.06 ml of 10(-3) infectivity titer of reovirus type I between the 2nd and 5th days of life. The pathological events consisted of necrotizing ependymitis, healing of the ependyma by gliovascular scarring, and obstruction of narrow bottlenecks such as the central canal. Histological findings were characterized by disorganization of the ependyma, formation of ependymal rosettes and microtubules, subependymal gliovascular scarring, and intracanalicular gliosis. These features are the same as those encountered clinically and provide strong evidence that stenosis of the central canal in man is a pathological lesion involving ependymal injury and scarring.

Teratogenic effects of neonatal arenavirus infection on the developing rat cerebellum are abrogated by passive immunotherapy.

The effects of viral infection on the developing nervous system and the potential of passive immunotherapy to protect against infection were examined. When 4-day-old Lewis rats were injected intracerebrally with lymphocytic choriomeningitis virus (LCMV) the majority of stem cells within the external granular layer of the developing cerebellum became infected. The infection progressed to the molecular layer, internal granular layer, and the Purkinje cells. By 15 days postinfection the molecular and internal granular layers of LCMV-infected cerebella were noticeably thinner than those in the controls and the individual folia were smaller. Neurons remained infected for up to 40 days as determined by immunohistochemistry. However, in rats treated with rat monoclonal anti-LCMV antibodies the staining was limited to the cells of ependyma and choroid plexus and was not detectable by 15 days postinfection. Macroscopically the infection resulted in pronounced hypoplasia, with the cerebella of 21-day-old LCMV-infected rats weighing 52 +/- 10 mg compared with 159 +/- 30 mg for control rats. Antibody-treated rats exhibited normal cerebellar size and development. Neutralizing antibodies specific for the viral GP-1 glycoprotein were protective but nucleoprotein-specific antibodies were not. Furthermore, suckling rat pups born of and nursed by LCMV-immune mothers were spared from cerebellar disease following neonatal infection. These results suggest that passive immunotherapy of neonates can provide effective protection against teratogenic effects of neonatal viral infection on the developing CNS.

The immune response in viral encephalitis.

The central nervous system (CNS) offers a unique organ system in which to study viral immunopathogenesis. The presence of the blood-brain barrier that restricts entry of cells and protein, the restricted expression of MHC antigens and the nonrenewable nature of the neuronal cell population offer challenges to the immune system for viral clearance and increase the chances for viral persistence. We have used Sindbis virus encephalitis in mice as a model system for the study of the development of immune reactions in the CNS and clearance of virus from neurons. The immune response to this and other viral infections of the CNS probably are initiated in peripheral lymphoid tissue followed by entry of activated T cells into the cerebrospinal fluid, meninges, and brain parenchyma. During Sindbis virus infection class I and II MHC antigens are expressed extensively on microglia which may present viral antigen produced by the infected neurons. Full development of the inflammatory response requires virus-specific T cells, but participating cells include NK cells, gamma delta T cells, monocytes and B cells. The entry of Ig-secreting B cells corresponds with the appearance of increased amounts of IgG and IgA in the cerebrospinal fluid. Clearance of Sindbis virus from the brain was studied using persistently infected severe combined immunodeficient (scid) mice. Passive transfer of immune serum or immune T cells to these infected mice demonstrated that antibody to a surface glycoprotein of the virus eliminated virus by a noncomplemented-mediated, noncytolytic mechanism. Immune T cells had no effect on virus replication.(ABSTRACT TRUNCATED AT 250 WORDS)

Postmitotic death is the fate of constitutively proliferating cells in the subependymal layer of the adult mouse brain.

The early development of the mammalian forebrain involves the massive proliferation of the ventricular zone cells lining the lateral ventricles. A remnant of this highly proliferative region persists into adult life, where it is known as the subependymal layer. We examined the proliferation kinetics and fates of the mitotically active cells in the subependyma of the adult mouse. The medial edge, the lateral edge, and the dorsolateral corner of the subependymal layer of the rostral portion of the lateral ventricle each contained mitotically active cells, but the dorsolateral region had the highest percentage of bromodeoxyuridine (BrdU)-labeled cells per unit area. Repeated injections of BrdU over 14 hr revealed a proliferation curve for the dorsolateral population with a growth fraction of 33%, indicating that 33% of the cells in this subependymal region make up the proliferating population. The total cell cycle time in this population was approximately 12.7 hr, with an S-phase of 4.2 hr. To examine the fate of these proliferating cells, we injected low concentrations of a replication-deficient, recombinant retrovirus directly into the lateral ventricles of adult mice for uptake by mitotically active subependymal cells. Regardless of the survival time postinjection (10 hr, 1 d, 2 d, or 8 d), the number of retrovirally labeled cells per clone remained the same (1 or 2 cells/clone). This suggests that one of the progeny from each cell division dies. Moreover, the clones remained confined to the subependyma and labeled cells were not seen in the surrounding brain tissue. Thus, while 33% of the dorsolateral subependymal cells continue to proliferate in adult life, the fate of the postmitotic progeny is death.

Detection of herpes simplex virus in the ependyma of experimentally infected mice.

Forty-five Swiss albino mice were inoculated with HSV-1 (strain McKrae) by the corneal route. The spread of HSV to the brain stem and the ventricular ependyma was investigated. The polymerase chain reaction (PCR) was used to detect HSV-DNA in the CSF during the course of infection. The ependyma of the third and fourth ventricle and the central canal contained viral antigen at a late stage of infection in up to 60% of mice. At this stage we found that many animals gave a positive PCR in the CSF although no antigen could be detected in the ependymal cells. The presence or absence of antigen containing cells could not be related to detection of HSV-DNA in the CSF. The results show that infection of the ventricular wall is not important for the spread of HSV to the CSF.

In vitro cytopathogenicity and in vivo virulence of two strains of canine parainfluenza virus.

In vivo and in vitro properties of two strains of canine parainfluenza virus (CPIV) were investigated. One strain, designated CPIV(+), induced syncytial giant cell formation and cytolysis in vitro, whereas the second strain, CPIV(-), caused only a mild strand-forming cytopathic effect with few, small syncytial giant cells. Vero cells infected with CPIV(+) or CPIV(-) were 100% positive for CPIV antigen as determined by immunofluorescent staining; however, 100% of CPIV(+) and less than 10% of CPIV(-) infected cells were hemadsorption positive. Sodium dodecyl sulphate-polyacrylamide gel electrophoresis analysis revealed no differences in electrophoretic mobility of viral polypeptides between both strains; however, in CPIV(-), reduced or absent synthesis of the putative HN and F1 proteins was observed. Isopycnic separation of CPIV(+) progeny virions showed a high proportion of viral particles with a buoyant density of 1.18 g/cm3. In contrast, CPIV(-) progeny virions had a heterogeneous density profile ranging from 1.08 to 1.18 g/cm3. Intracerebral infection of six ferrets with CPIV(+) resulted in moderate lymphocytic and histiocytic choroiditis, meningitis, and ependymitis, whereas CPIV(-) infection caused only mild to moderate inflammation. Immunohistologically, CPIV antigen was prominent in ependymal lining cells of the ventricles in CPIV(+)-infected ferrets and was reduced or lacking in CPIV(-)-infected ferrets (n = 6). Sham-injected ferrets (n = 6) did not have histologic lesions and no viral antigen was identified. The present findings suggest that certain changes in the activities of CPIV glycoproteins may lead to alterations of CPIV virulence in vivo.

Borna disease virus replicates in astrocytes, Schwann cells and ependymal cells in persistently infected rats: location of viral genomic and messenger RNAs by in situ hybridization.

Borna disease (BD) is an immune-mediated neurological disease caused by infection of the nervous system with a negative strand RNA virus, Borna disease virus (BDV). The host range for BDV is broad and extends from birds to primates. A BDV-like agent may cause disease in humans. Until recently, BDV-infected neural cells could only be identified immunocytochemically using serum from BDV-infected animals. The advent of BDV cDNA clones allowed definition of the relationship between viral nucleic acids and viral proteins in vivo. In situ hybridization with strand-specific RNA probes from a BDV cDNA clone, pAF4, identified BDV genomic RNA and BDV mRNAs in neurons, astrocytes, Schwann cells and ependymal cells in an anatomic distribution consistent with that of BDV proteins. Genomic RNA was contained primarily within the nucleus, whereas mRNAs were found in both the nuclear and cytoplasmic compartments. Viral RNAs were demonstrated in neurons expressing BDV proteins and in glial cells by combined techniques of immunocytochemistry and in situ hybridization.

The pathogenesis of spinal cord involvement in the encephalomyelitis of mice caused by neuroadapted Sindbis virus infection.

Weanling mice develop an acute encephalomyelitis with high mortality after intracerebral inoculation of neuroadapted Sindbis virus. The mice develop kyphoscoliosis and hindlimb paralysis. Immunohistochemical and in situ hybridization studies have demonstrated virus in the gray matter of the brain and spinal cord. Ventral horn cells are prominently infected, providing an anatomical basis for the clinical poliomyelitis. A novel route of spread of inoculated virus within the central nervous system has been found. The virus enters the ventricular system, and then travels caudally in the central canal of the spinal cord where ependymal cells are infected. The virus subsequently spreads into the gray matter. The distribution of virus in the spinal cord is likely dependent both on variations in the susceptibility of neural cells and on this route of entry and subsequent spread.

Selective tropism of a neurotropic coronavirus for ependymal cells, neurons, and meningeal cells.

The ability of a neurotropic virus, mouse hepatitis virus type 3 (MHV3), to invade the central nervous system (CNS) and to recognize cells selectively within the brain was investigated in vivo and in vitro. In vivo, MHV3 induced in C3H mice a genetically controlled infection of meningeal cells, ependymal cells, and neurons. In vitro, purified MHV3 bound to the surface of isolated ependymal cells and cultured cortical neurons but not to oligodendrocytes or cultured astrocytes. MHV3 replicated within cultured cortical neurons and neuroblastoma cells (NIE 115); infected cultured neurons nonetheless survived and matured normally for a 7-day period postinfection. On the other hand, MHV3 had a low affinity for cortical glial cells or glioma cells (C6 line), both of which appear to be morphologically unaltered by viral infection. Finally, MHV3 infected and disrupted cultured meningeal cells. This suggests that differences in the affinity of cells for MHV3 are determinants of the selective vulnerability of cellular subpopulations within the CNS. In vivo, a higher titer of virus was needed for CNS penetration in the genetically resistant (A/Jx) mice than in the susceptible (C57/BL6) mouse strain. However, in spite of viral invasion, no neuropathological lesions developed. In vitro viral binding to adult ependymal cells of susceptible and resistant strains of mice was identical. Genetic resistance to MHV3-CNS infection appeared to be mediated both by a peripheral mechanism limiting viral penetration into the CNS and by intra-CNS mechanisms, presumably at a stage after viral attachment to target cells.

Mycoplasma pneumoniae-induced hydrocephalus in hamsters.

Hydrocephalus was induced in neonatal hamsters after intracerebral inoculation of Mycoplasma pneumoniae. Examination of the ependyma from affected animals by electron microscopy did not reveal mycoplasma. However, in an ependymal organ culture system, M. pneumoniae cytadsorbed to ependymal cells.

Antibodies to a scrapie prion protein.

Scrapie is a slow infection of the nervous system which progresses in the absence of any apparent immune response. The recent development of a large-scale purification protocol for scrapie prions made it possible to obtain substantial quantities of electrophoretically purified prion protein (PrP 27-30) and we report here on the successful production of a rabbit antiserum to PrP 27-30. The antiserum reacted with PrP 27-30 and several lower molecular weight proteins as shown by Western blots; it did not react with protein preparations from uninfected brains. Discrete structures in the subependymal region of scrapie-infected hamster brains were stained immunocytochemically. These same structures also stained with Congo red dye and showed green birefringence with polarized light, a characteristic of purified prion rods. This staining pattern suggests that they are amyloid plaques.

Asymmetric budding of viruses in ependymal and choroid plexus epithelial cells.

Sendai virus injected intracerebrally into 3-week-old mice caused infection of ependymal and choroid plexus epithelial cells. Budding of mature viruses occurred only from the apical surfaces of these cells. The viral peplomere proteins, haemagglutinin-neuraminidase and fusion, were concentrated on the apical portion of the ependymal cells, while the nucleocapsid-associated polymerase protein was dispersed throughout the cytoplasm. This indicates that intracellular routing of the virus envelope glycoproteins to the cell surface may be one of the factors that determines the site of virus budding. Vesicular stomatitis and vaccinia viruses, on the other hand, budded or egressed predominantly from the baso-lateral cell surfaces.

Viral receptors on isolated murine and human ependymal cells.

Viruses that infect ependyma cause ependymitis in humans and hydrocephalus in experimental animals. We report that reovirus type 1 (which induces hydrocephalus in mice) binds to the surface of isolated human and murine ciliated ependymal cells. With the use of recombinant viral clones, the binding property was mapped to the type 1 viral hemagglutinin, which also determines in vivo the affinity of reovirus type 1 for ependyma. Mumps virus, measles virus, parainfluenza type 3, and herpes simplex virus type 1 bind to murine ependyma cells, whereas reovirus type 3, herpes simplex virus type 2, and poliovirus type 2 do not.

Cytadsorption of Mycoplasma pulmonis to rat ependyma.

Mycoplasma pulmonis was inoculated intracerebrally into neonatal rats. Hydrocephalus was induced, and the lateral ventricles and aqueduct were examined by scanning electron microscopy. Mycoplasmas were observed to be cytadsorbed to the ependymal surface.