Evolution of mouse hepatitis virus (MHV) during chronic infection: quasispecies nature of the persisting MHV RNA.

Coronavirus infection of mice has been used extensively as a model for the study of acute encephalitis and chronic demyelination. To examine the evolution of coronavirus RNA during chronic demyelinating infection, we isolated RNA from intracerebrally inoculated mice at 4, 6, 8, 13, 20, and 42 days postinfection and used reverse transcription-polymerase chain reaction amplification methods (RT-PCR) to detect viral sequences. RNA sequences from two viral structural genes, the spike gene and the nucleocapsid gene, were detected throughout the chronic infection. In contrast, infectious virus was not detectable from brain homongenates beyond 13 days postinfection. These results indicate that coronavirus RNA persists in the brain at times when infectious virus is not detected. To determine if genetic changes were occurring during viral replication in the host, we cloned and sequenced the RT-PCR products from the spike and nucleocapsid regions and analyzed the sequences for mutations. Sequencing of the cloned products revealed that a variety of mutant forms of viral RNA persisted in the CNS, including point mutants, deletion mutants, and termination mutants. The mutations accumulated during persistent infection in both the spike and the nucleocapsid sequences, with greater than 65% of the mutations encoding amino acid changes. These results show that a diverse population or quasispecies consisting of mutant and deletion variant viral RNAs (which may not be capable of producing infectious virus particles) persists in the central nervous system of mice during chronic demyelinating infection. The implications of these results for the role of persistent viral genetic information in the pathogenesis of chronic demyelination are discussed.

Mutations associated with viral sequences isolated from mice persistently infected with MHV-JHM.

Mouse hepatitis virus JHM (JHMV or MHV-4) induces subacute and chronic demyelination in rodents and has been studied as a model human demyelinating diseases, such a multiple sclerosis. However, despite intensive investigation, the state of JHMV during chronic disease is poorly understood. Using reverse transcription-polymerase chain reaction amplification (RT-PCR) to "rescue" viral RNA, we have found that JHMV-specific sequences persist for at least 787 days after intracerebral inoculation of experimental mice. Analysis of persisting viral RNA reveals that it is extensively mutated, and we hypothesize that the mutations observed reflect adaptation of the viral quasispecies to low-level intracellular replication during chronic disease.

Nasal pyriform aperture stenosis and the holoprosencephaly spectrum.

Recent reports have described congenital nasal pyriform aperture stenosis, but do not address its etiology in detail. We describe a child with nasal pyriform aperture stenosis, submucus cleft palate, and hypoplastic maxillary sinuses. Chromosome analysis revealed a ring chromosome 18. Awareness of the association of midline facial defects with midline brain defects allowed us to predict that features of the holoprosencephaly sequence would be found. Subsequent evaluation revealed growth hormone deficit. Eventually the child manifested a single central incisor. We review the association between midline facial defects and holoprosencephaly to remind the otolaryngologist of the need to look at the whole patient as he treats specific upper airway problems.

The terminally ill patient on an acute psychiatric unit. A case study.

1. As mental health professionals, we frequently fail to realize that we are subject to the same emotions and coping mechanisms as the lay community. 2. At times it seems safer to focus on a patient's medical condition than to deal with difficult, unresolved psychological issues. 3. By using existing resources within our institution, our interdisciplinary team faced many of their unresolved issues about death, such as countertransference, and were able to then help our patient through his final stage of life.

Hirschsprung disease: etiologic implications of unsuccessful prenatal diagnosis.

We describe an infant with Hirschsprung disease (congenital aganglionosis of the intestine) involving the colon and terminal ileum. Midtrimester prenatal diagnosis of this disorder in this infant was attempted utilizing amniotic fluid disaccharidase analyses, ultrasound, and amniography. Decreased disaccharidase activities in amniotic fluid have been reported previously in association with other forms of intestinal obstruction. At 15 weeks' gestation, normal amniotic fluid disaccharidase levels were obtained. Serial ultrasound evaluations did not indicate any pathology, and the results from amniography were inconclusive. The implication of the normal disaccharidase values is that Hirschsprung disease may in some cases result from degeneration of intestinal ganglia after 16 weeks' gestation rather than from faulty migration of neural crest cells. The inheritance of Hirschsprung disease is generally consistent with sex-modified multifactorial inheritance with a lower threshold of expression in males. The case we report has a family history of three affected first- and second-degree relatives. Autosomal dominance with variable expressivity is a possible explanation in this family.

Placental cultures for cytogenetic assessment in saline-aborted fetuses.

Confirmation of cytogenetically abnormal fetuses following saline abortion has been shown to be possible with the placenta as the source of viable fetal cells. The method is described in detail. In one third of cultures, only female cells were present. Differentiation between maternal and female fetal tissue when no numerical or structural cytogenetic disorder is present requires detailed analysis of fluorescently stained chromosomes for polymorphisms.

Males with chronic myeloid leukemia and the 45, XO, Ph1 chromosome pattern.

Two cases of chronic myeloid leukemia in which the cytogenetic clone was 45, XO, Ph(1) are described and compared with 20 cases recorded in the literature. The 45, XO line is peculiar to the leukemic cells and is not a manifestation of constitutional mosaicism. It probably arises from a 46, XY, Ph(1) line by loss of the Y chromosome. Because of the few cases reported, one cannot be certain that these men have a better prognosis, although the median survival time suggests that this is so. Infertility is not part of this disorder.