Isolation of a novel adenovirus from Rousettus leschenaultii bats from India.

Surveillance work was initiated to study the presence of highly infectious diseases like Ebola-Reston, Marburg, Nipah and other possible viruses that are known to be found in the bat species and responsible for causing diseases in humans. A novel adenovirus was isolated from a common species of fruit bat (Rousettus leschenaultii) captured in Maharashtra State, India. Partial sequence analysis of the DNA polymerase gene shows this isolate to be a newly recognized member of the genus Mastadenovirus (family Adenoviridae), approximately 20% divergent at the nucleotide level from Japanese BatAdV, its closest known relative.

Child injury in a changing world.

The importance of child injuries has now been recognised as a significant public health problem internationally. The World Health Organisation (WHO) and United Nations Children's Fund (UNICEF) have recently published the first world report on child injury prevention. As infectious diseases decline, the relative importance of injury has increased, but the pace of change of global processes means that absolute increases in injury may occur over the next 20-30 years. This paper examines child injury in a changing world by outlining the ways in which the forces of globalisation, urbanisation, motorisation and environmental change could have an impact on injury epidemiology and policy. We consider how those in public health and those in the injury field should respond to the changing world of injury. Child injury prevention needs to be incorporated into planning for the rapidly changing urban environments of low-income countries and strategies devised for the large numbers of people displaced by environmental change.

The hazards of daily life: an historical perspective on adult unintentional injuries.

Unintentional injuries are a major public health problem. This paper analyses coroners' inquests from Sussex, England, for the period 1485-1688 to consider the circumstances surrounding adult unintentional injury deaths. Parallels with the situation today are examined. Travel was found to be the most hazardous activity, drowning was also highly significant and there were large differences between men and women.

UNICEF's child injury league table. An analysis of legislation: more mixed messages.

This paper presents a summary table and discussion of legislation related to child injury prevention in member countries of the Organisation for Economic Cooperation and Development. The table is an expanded version of the one which appeared in the UNICEF Report Card "Child Deaths by Injury in Rich Countries" (2001). A commentary is provided on the variations in legislation between countries in terms of range and form of measures and an estimate of degree of enforcement. As legislation is generally considered a powerful tool in injury prevention, the paper examines whether those countries with the widest range of legislation and the strongest enforcement have made the most progress in reducing child injury deaths since the 1970s. It also considers whether a commitment to extensive legislation is reflected in a country's position in the UNICEF league table of injury death. The initial conclusion to these two basic issues is that no clear picture can be seen and we thus need to know far more about the relationship between legislation and societies and cultures as they vary from place to place. This paper hopes to stimulate more widespread debate about the role of legislation in different countries.

Rescue of defective poliovirus RNA replication by 3AB-containing precursor polyproteins.

This study demonstrates the in vitro complementation of an RNA replication-defective lesion in poliovirus RNA by providing a replicase/polymerase precursor polypeptide [P3(wt) (wild type)] in trans. The replication-defective mutation was a phenylalanine-to-histidine change (F69H) in the hydrophobic domain of the membrane-associated viral protein 3AB. RNAs encoding wild-type forms of protein 3AB or the P3 precursor polypeptide were cotranslated with full-length poliovirus RNAs containing the F69H mutation in a HeLa cell-free translation/replication assay in an attempt to trans complement the RNA replication defect exhibited by the 3AB(F69H) lesion. Unexpectedly, generation of 3AB(wt) in trans was not able to efficiently complement the defective replication complex; however, cotranslation of the large P3(wt) precursor protein allowed rescue of RNA replication. Furthermore, P3 proteins harboring mutations that resulted in either an inactive polymerase or an inactive proteinase domain displayed differential abilities to trans complement the RNA replication defect. Our results indicate that replication proteins like 3AB may need to be delivered to the poliovirus replication complex in the form of a larger 3AB-containing protein precursor prior to complex assembly rather than as the mature viral cleavage product.

Utilization of a mammalian cell-based RNA binding assay to characterize the RNA binding properties of picornavirus 3C proteinases.

Using an assay capable of detecting sequence-specific RNA/protein interactions in mammalian cells, we demonstrate that the poliovirus and rhinovirus 3C proteinases are able to bind structured target RNA sequences derived from their respective 5' noncoding regions in vivo. Specific RNA binding by poliovirus 3C was found to be dependent on the integrity of stem-loop d of the RNA cloverleaf structure located at the 5' end of poliovirus genomic RNA. In contrast, mutation of stem-loop b did not prevent this in vivo interaction. However, mutation of stem-loop b, which serves as the RNA binding site for a cellular co-factor important for efficient poliovirus replication, did significantly attenuate the efficiency of 3C RNA binding in vivo and 3CD RNA binding in vitro. This in vivo protein:RNA binding assay was also used to identify several residues in 3C that are critical for RNA binding, but dispensable for 3C proteinase activity. The mammalian cell-based RNA binding assay described in this study may have considerable potential utility in the future detection or analysis of in vivo RNA/protein interactions unrelated to the 3C/RNA interaction described here.

Poly (rC) binding protein 2 forms a ternary complex with the 5'-terminal sequences of poliovirus RNA and the viral 3CD proteinase.

Poly(rC) binding protein 2 (PCBP2) forms a specific ribonucleoprotein (RNP) complex with the 5'-terminal sequences of poliovirus genomic RNA, as determined by electrophoretic mobility shift assay. Mutational analysis showed that binding requires the wild-type nucleotide sequence at positions 20-25. This sequence is predicted to localize to a specific stem-loop within a cloverleaf-like secondary structure element at the 5'-terminus of the viral RNA. Addition of purified poliovirus 3CD to the PCBP2/RNA binding reaction results in the formation of a ternary complex, whose electrophoretic mobility is further retarded. These properties are consistent with those described for the unidentified cellular protein in the RNP complex described by Andino et al. (Andino R, Rieckhof GE, Achacoso PL, Baltimore D, 1993, EMBO J 12:3587-3598). Dicistronic RNAs containing mutations in the 5' cloverleaf-like structure of poliovirus that abate PCBP2 binding show a decrease in RNA replication and translation of gene products directed by the poliovirus 5' noncoding region in vitro, suggesting that the interaction of PCBP2 with these sequences performs a dual role in the virus life cycle by facilitating both viral protein synthesis and initiation of viral RNA synthesis.

Replication-competent picornaviruses with complete genomic RNA 3' noncoding region deletions.

The genomic RNA 3' noncoding region is believed to be a major cis-acting molecular genetic determinant for regulating picornavirus negative-strand RNA synthesis by promoting replication complex recognition. We report the replication of two picornavirus RNAs harboring complete deletions of the genomic RNA 3' noncoding regions. Our results suggest that while specific 3'-terminal RNA sequences and/or secondary structures may have evolved to promote or regulate negative-strand RNA synthesis, the basic mechanism of replication initiation is not strictly template specific and may rely primarily upon the proximity of newly translated viral replication proteins to the 3' terminus of template RNAs within tight membranous replication complexes.

Requirement of poly(rC) binding protein 2 for translation of poliovirus RNA.

Poly(rC) binding protein 2 (PCBP2) is one of several cellular proteins that interact specifically with a major stem-loop domain in the poliovirus internal ribosome entry site. HeLa cell extracts subjected to stem-loop IV RNA affinity chromatography were depleted of all detectable PCBP2. Such extracts were unable to efficiently translate poliovirus RNA, although extracts recovered from control columns of matrix unlinked to RNA retained full translation activity. Both translation and production of infectious progeny virus were restored in the PCBP2-depleted extracts by addition of recombinant PCBP2, but not by PCBP1, which is a closely related member of the protein family. The data show that PCBP2 is an essential factor, which is required for efficient translation of poliovirus RNA in HeLa cells.

Translation and replication properties of the human rhinovirus genome in vivo and in vitro.

The poor translation efficiency of genome-length human rhinovirus RNA in vitro using HeLa cell extract-supplemented rabbit reticulocyte lysate has hampered the study of rhinovirus IRES-mediated translation and polyprotein synthesis in a cell-free system. In contrast, the efficient in vitro translation characteristics of poliovirus RNAs have ultimately allowed the programming of cell-free coupled translation/replication extracts which are able to produce infectious poliovirus particles in vitro. A possible explanation for the decreased burst size observed during the course of a rhinovirus infection, compared to poliovirus infection, is reduced levels of polyprotein synthesis in vivo. In order to test this hypothesis and extend in vitro translation/replication technology to the study of human rhinoviruses, a chimeric cDNA construct was engineered which allowed the in vitro synthesis of T7 transcripts containing the intact poliovirus type 1 (PV1) 5' noncoding region (5' NCR) and initiation codon upstream of the human rhinovirus 14 (HRV14) polyprotein-coding region and 3'-terminal sequences. These chimeric RNAs translated efficiently in vitro and were used successfully to program a cell-free replication extract. Unexpectedly, parental HRV14 RNAs also translated efficiently in the HeLa cell-free translation/replication extract but replicated less efficiently than the chimera in vitro. The chimeric HRV14/PV1 RNAs were infectious and gave rise to a virus with a growth phenotype similar to that of parental HRV14. Preliminary characterization of this chimeric virus suggests that the biological properties characteristic of rhinovirus in vivo are determined primarily by the rhinovirus gene products. Although the translation efficiency of the HRV14 5' NCR may be a limitation in rabbit reticulocyte lysate-based in vitro translation extracts, it does not appear to be a major limiting determinant for growth of rhinovirus in vivo or replication in the HeLa cell-free extract.

Determinants of membrane association for poliovirus protein 3AB.

Poliovirus protein 3AB may serve as the lipophilic carrier of a protein primer (VPg or 3B) used for the initiation of genomic viral RNA synthesis. In order to study the membrane-protein interactions of 3AB required for its role in poliovirus RNA replication, we have developed an in vitro membrane association assay capable of distinguishing membrane-bound from non-membrane-bound proteins that are cotranslated together in the presence of canine microsomal membranes. This assay utilizes equilibrium sedimentation analysis in high density sucrose gradients to measure membrane association of both wild type and mutated forms of 3AB. Using this assay and other biochemical assays, we have identified the following properties of the 3AB-membrane interaction: (a) 3AB is able to post-translationally associate with microsomal membranes, (b) 3AB is able to associate with membranes in a manner consistent with that of an integral membrane protein, (c) 3AB contains a critical hydrophobic sequence within the carboxyl-terminal half of the protein that is required for membrane association, and (d) the introduction of charged residues into this hydrophobic sequence disrupts the 3AB membrane-protein interaction. Taken together, these studies indicate that poliovirus protein 3AB associates tightly with biological membranes de novo in a manner that would allow it to serve as a lipophilic anchor for the assembly of the poliovirus RNA replication complex.

Ectrodactyly and proximal/intermediate interstitial deletion 7q.

We report on an individual with severe mental retardation, seizures, microcephaly, unusual face, scoliosis, and cleft feet and cleft right hand. The chromosomal study showed a proximal interstitial deletion 7q (q11.23q22). From our review of the literature, 11 patients have been reported with ectrodactyly (split hand/split foot malformation) and proximal/intermediate interstitial deletions or rearrangements of 7q. The critical segment for ectrodactyly seems to be located between 7q21.2 and 7q22.1. This malformation is present in 41% of the patients whose deletion involves the critical segment.

Role of eicosanoids in human and experimental colitis.

Prostaglandins, thromboxanes, and leukotrienes (collectively called eicosanoids) are increased at sites of inflammation and contribute to the manifestations of inflammation, such as hyperemia, hyperalgesia, edema, and inflammatory cell infiltration. Inhibition of eicosanoid production is the basic mechanism of action of corticosteroids and of nonsteroidal antiinflammatory drugs. Eicosanoid synthesis is also increased in human and experimental inflammatory bowel disease. Leukotriene B4 is the most potent proinflammatory eicosanoid, and in vivo production of this compound is the predominant eicosanoid in colitis. Recent experimental data demonstrate that selective inhibition of leukotrienes may be a therapeutic strategy to reduce inflammation in inflammatory bowel disease.

Indomethacin and ibuprofen enhance anoxia-induced hyperemia in rat brain.

Cerebral blood flow (CBF) in the rat was monitored by a venous outflow technique with an extracorporeal circulation, which allows for the continuous recording of CBF over periods of several hours. Following intraperitoneal administration, two non-steroidal anti-inflammatory agents, indomethacin and ibuprofen (0.01 and 0.1 mg/kg) showed a tendency to increase resting CBF and the reactive hyperemia elicited by a brief (24 s) anoxic challenge was potentiated. A third agent, diclofenac sodium, was less effective, enhancing the hyperemic response only at the higher (0.1 mg/kg) dose level. The results indicate that indomethacin and ibuprofen may be of value in the treatment of diseases involving cerebrovascular insufficiency.

The effects of nifedipine and felodipine on cerebral blood flow during anoxic episodes.

Cerebral blood flow (CBF) in the rat was monitored by a venous outflow technique with an extracorporeal circulation, which allows for the continuous recording of flow over periods of several hours. Brief periods of anoxia increase the rate of flow. The dihydropyridine calcium antagonists did not affect basal flow rate and depressed the increase in CBF elicited by anoxia. These findings may have significant implications for the therapeutic use of dihydropyridine calcium antagonists in brain ischaemia.

Naloxone enhances cerebral reactive hyperemia in the rat.

Cerebral blood flow (CBF) in the rat was monitored by a venous outflow technique with an extracorporeal circulation, which allows for the continuous recording of CBF over several hours. Morphine and the opiate antagonist, naloxone, were tested for their effects on the reactive hyperemia that follows a brief anoxic challenge. Morphine (5.0 mg/kg) significantly reduced the peak increase in flow during the hyperemia and, at both of the doses used (1.0 and 5.0 mg/kg), caused a small, nonsignificant increase in the duration of the reactive hyperemia. Naloxone (0.1 and 1.0 mg/kg) enhanced basal CBF rates and significantly prolonged the duration of the reactive hyperemia. These effects of naloxone may account for its beneficial effects in the treatment of cerebral ischemia.

Adenosine deaminase inhibitors enhance cerebral anoxic hyperemia in the rat.

Cerebral blood flow in the rat was monitored by a venous outflow technique with an extracorporeal circulation, which allows for the continuous recording of flow over periods of several hours. The adenosine deaminase inhibitors erythro-9-(2-hydroxy-3-nonyl)adenine (EHNA) (1.0-100 micrograms/kg) and deoxycoformycin (0.1-1 micrograms/kg) potentiated the reactive hyperemia elicited by a brief (24-s) anoxic challenge. Basal flow rate was unaltered by EHNA administration and slightly enhanced by deoxycoformycin. The results are consistent with the hypothesis that adenosine plays a significant role in cerebral vascular regulation and suggest that low doses of these deaminase inhibitors may be useful in the treatment of cerebral vascular insufficiency.

The effects of lidoflazine and flunarizine on cerebral reactive hyperemia.

Cerebral blood flow in the rat was monitored by a venous outflow technique with an extracorporeal circulation, which allows for the continuous recording of flow over periods of several hours. The bi-fluorophenyl-piperazine derivatives, lidoflazine and flunarizine, enhanced the reactive hyperemia elicited by a brief (30 s) anoxic challenge. They did not alter resting cerebral blood flow rates. Verapamil, a potent calcium slow channel blocker, decreased resting flow rates but did not alter the duration of the reactive hyperemia. As lidoflazine and flunarizine are potent inhibitors of adenosine uptake, whereas verapamil is not, the results are consistent with the hypothesis that adenosine plays a significant role in cerebral vascular autoregulation.

Additional manifestations of the Neu-Laxova syndrome.

A newborn female with intrauterine growth retardation, bilateral cleft lip and palate, absent external nares and eyelids, low set ears, short contracted limbs, webbed digits, intestinal malrotation, and unilateral renal agenesis is reported. These multiple malformations are considered part of the Neu-Laxova syndrome.