The next generation: Pregnancy in adolescents and women living with perinatally acquired HIV in South Africa.

BACKGROUND: An increasing number of girls living with perinatally acquired HIV (PHIV) are reaching adolescence and adulthood and becoming pregnant. Youth living with PHIV (YLPHIV) may have HIV-associated infections/complications, long-term exposure to antiretroviral treatment (ART), drug resistance and increased psychosocial challenges, which may adversely affect pregnancy outcomes. There is a lack of published studies on pregnancy in YLPHIV in sub-Saharan Africa. Objectives. To describe characteristics of pregnant South African (SA) YLPHIV and their pregnancy outcomes. METHODS: We retrospectively identified pregnancies in YLPHIV, who were diagnosed with HIV when they were <12 years old and before their first pregnancy (as a proxy for perinatal route of infection), from routinely collected data in Western Cape Province, SA (2007 - 2018). We combined these with pregnancies from a Johannesburg cohort of YLPHIV. Results. We identified 258 pregnancies among 232 females living with likely PHIV; 38.8% of pregnancies occurred in YLPHIV ≤16 years old, 39.1% at age 17 - 19 years and 22.1% at age ≥20 years. In recent years, a steady increase in the number of pregnancies in YLPHIV was noted; more than two-thirds occurred during 2016 - 2018. ART was commenced prior to pregnancy in 84.9% of YLPHIV, during pregnancy in 6.6% and was not commenced by pregnancy end date in 8.5%. Of the pregnancies in young women with documented outcomes (88.8%; n=229), 80.3% were live births, 14.4% terminations, 3.1% miscarriages and 2.2% stillbirths. Mother-to-child transmission of HIV occurred in 2.2% of infants, 75.3% were uninfected when last tested and 22.6% had unknown HIV status. Among YLPHIV with CD4 counts available within 12 months of pregnancy end date (n=202), 20.3% had a CD4 count <200 cells/µL, 43.1% CD4 count 200 - 499 cells/µL and 36.6% CD4 count ≥500 cells/µL. Among those with a viral load (VL) available within 12 months of pregnancy end date (n=219), 66.7% had a VL <400 copies/mL, 5.0% VL 400 - 999 copies/mL and 28.3% VL ≥1 000 copies/mL. Of 186 neonates, 20.4% were preterm deliveries (<37 weeks' gestation). Among neonates with known birthweight (n=176), the mean birthweight was 2 900 g (95% confidence interval (CI) 2 747 - 2 935 g) and 20.5% had a low birthweight (<2 500 g). One congenital malformation (musculoskeletal) and 2 neonatal deaths were recorded. CONCLUSIONS: In recent years, the number of pregnancies in YLPHIV has increased. A considerable proportion of pregnancies occurred in YLPHIV ≤16 years old. A high proportion of pregnancies was electively terminated. The prevalence of elevated VL and poor immunological status among pregnant YLPHIV is concerning.

Comparative analysis of the Metarhizium anisopliae secretome in response to exposure to the greyback cane grub and grub cuticles.

Metarhizium anisopliae is a well-characterized biocontrol agent of a wide range of insects including cane grubs. In this study, a two-dimensional (2D) electrophoresis was used to display secreted proteins of M. anisopliae strain FI-1045 growing on the whole greyback cane grubs and their isolated cuticles. Hydrolytic enzymes secreted by M. anisopliae play a key role in insect cuticle-degradation and initiation of the infection process. We have identified all the 101 protein spots displayed by cross-species identification (CSI) from the fungal kingdom. Among the identified proteins were 64-kDa serine carboxypeptidase, 1,3 beta-exoglucanase, Dynamin GTPase, THZ kinase, calcineurin like phosphoesterase, and phosphatidylinositol kinase secreted by M. ansiopliae (FI-1045) in response to exposure to the greyback cane grubs and their isolated cuticles. These proteins have not been previously identified from the culture supernatant of M. anisopliae during infection. To our knowledge, this the first proteomic map established to study the extracellular proteins secreted by M. ansiopliae (FI-1045) during infection of greyback cane grubs and its cuticles.

Admissions processes at the seven United Kingdom veterinary schools: a review.

The major challenge in veterinary undergraduate admissions is to select those students with most suitability for veterinary training and careers from a large and diverse pool of applicants with very high academic ability. This paper describes a review of the admissions processes of the seven veterinary schools in the UK. There was significant commonality in the entry requirements and the criteria upon which the schools made decisions on candidates. There was some variation in the procedures used by individual schools to select candidates, but common themes existed within these processes. All of the schools evaluated both academic and non-academic factors for individual applicants, and all used interviews in some format as a selection tool after an initial short-listing process. The procedures and approaches to selection processes are compared and discussed.

Low-dose nonenhanced head CT protocol for follow-up evaluation of children with ventriculoperitoneal shunt: reduction of radiation and effect on image quality.

BACKGROUND AND PURPOSE: Children with a shunt for hydrocephalus often undergo multiple follow-up head CT scans, increasing the risk for long-term effects of ionizing radiation. The purpose of our study was to evaluate if an unenhanced low-dose head CT could consistently provide acceptable image quality and diagnostic information. MATERIALS AND METHODS: Ninety-two children (mean age, 9 years; range, 8 months to 21 years; 45 boys and 47 girls) with a shunt for hydrocephalus and no clinical evidence of shunt malfunction who were referred for a follow-up nonenhanced head CT were included in the study. All studies were performed on a 4-section multidetector CT. Two CT studies were selected retrospectively for each patient, 1 performed at standard dose (220 mA) and 1 at low dose (80 mAs). Two radiologists independently evaluated and graded both standard-dose and low-dose studies for various image quality parameters. Attenuation and noise levels were measured, and gray-white differentiation and contrast-to-noise ratio (CNR) were calculated. RESULTS: Low-dose CT resulted in 63% mean dose reduction. All low-dose CT scans were diagnostically acceptable. Image quality parameters were significantly lower at low dose (P = .0001) except for the parameters for streak artifacts (P = .46) and need for further imaging (P = .47), which were higher. Mean noise levels were significantly higher (P = .001) in low-dose studies, whereas CNR was significantly higher in standard dose CT (P = .001). A moderate to perfect agreement was noted between the 2 readers with regard to image quality assessment (65%-99%). CONCLUSION: Low-dose nonenhanced head CT consistently provides diagnostically acceptable images with relevant diagnostic information in children with VP shunts resulting in substantial dose savings.

Yellow leaf of sugarcane is caused by at least three different genotypes of sugarcane yellow leaf virus, one of which predominates on the Island of Réunion.

The genetic diversity of sugarcane yellow leaf virus (SCYLV) was analyzed with 43 virus isolates from Réunion Island and 17 isolates from world-wide locations. We attempted to amplify by reverse-transcription polymerase chain reaction (RT-PCR), clone, and sequence four different fragments covering 72% of the genome of these virus isolates. The number of amplified isolates and useful sequence information varied according to each fragment, whereas an amplicon was obtained with diagnostic primers for 59 out of 60 isolates (98%). Phylogenetic analyses of the sequences determined here and additional sequences of 11 other SCYLV isolates available from GenBank showed that SCYLV isolates were distributed in different phylogenetic groups or belonged to single genotypes. The majority of isolates from Réunion Island were grouped in phylogenetic clusters that did not contain any isolates from other origins. The complete six ORFs (5612 bp) of five SCYLV isolates (two from Réunion Island, one from Brazil, one from China, and one from Peru) were amplified, cloned, and sequenced. The existence of at least three distinct genotypes of SCYLV was shown by phylogenetic analysis of the sequences of these isolates and additional published sequences of three SCYLV isolates (GenBank accessions). The biological significance of these genotypes and of the origin of the distinct lineage of SCYLV in Réunion Island remains to be determined.

A variable region of the sugarcane bacilliform virus (SCBV) genome can be used to generate promoters for transgene expression in sugarcane.

Four promoters derived from sugarcane bacilliform virus (SCBV) were compared and characterised. Three were obtained by PCR amplification of purified virion DNA extracted from three sugarcane cultivars. The fourth promoter was obtained by subcloning from an almost genome-length clone of SCBV. All promoters were able to drive stable expression of beta-glucuronidase in sugarcane. The PCR-derived promoter sequences shared more DNA homology with banana streak virus than to the subcloned SCBV. The subcloned promoter was the strongest expressing and was able to drive reporter gene expression in vitro and in the leaves, meristems and roots of glasshouse-grown sugarcane. Expression levels were at least equal to or higher than those measured for the maize polyubiquitin promoter.

Sugarcane bacilliform virus encapsidates genome concatamers and does not appear to integrate into the Saccharum officinarum genome.

Sugarcane bacilliform virus (SCBV) DNA molecules larger than the complete genome length of 7.6 kbp were detected in infected plants and in virions. We have confirmed that these high molecular weight nucleic acids were open circular DNA and viral in origin. Due to their open circular conformation, accurate size determination of the DNA molecules was not possible using conventional electrophoresis. Using field inversion gel electrophoresis (FIGE), however, the DNA appeared to increase in genome size increments, with sizes ranging from 1 to 4 genomes (31 kbp) detected. The DNA was packaged into virions, which may explain the observation of purified virions with lengths corresponding to one, two or three times the modal length of 130 nm. The DNA products were possibly concatamers formed during replication as a result of a terminal overlap on the sense strand, and were shown to be overlapped individual genome-length molecules and not covalently-bonded continuous DNA strands. Southern analysis indicated that SCBV sequences are not integrated into the sugarcane genome and that the high molecular weight DNA observed in the sugarcane accessions analysed represents SCBV concatamers.

Sequence analysis of an Australian isolate of sugarcane bacilliform badnavirus.

The genome of an Australian isolate of Sugarcane bacilliform virus (SCBV-IM) was cloned, sequenced and analysed. The genome consisted of 7687 nucleotides and contained three open reading frames which were similar in size and organisation to those of other badnaviruses. SCBV-IM was found to be most similar to the SCBV-Morocco isolate with amino acid sequence similarity of 91.4 %, 83.8 % and 85.3 % in the ORF I, II and III coding regions, respectively. Phylogenetic analysis of the SCBV-IM ORF III deduced amino acid sequence showed that SCBV isolates were more closely related to each other than to other badnaviruses. Amplification of SCBV sequences from three different sugarcane varieties revealed considerable variability in the viral populations, both within single infected plants as well as between infected plants, suggesting that the SCBV isolates sequenced to date may not be representative of the range of virus variability.

Cyclin D1 expression in transitional cell carcinoma of the bladder: correlation with p53, waf1, pRb and Ki67.

Normal cell proliferation is closely regulated by proteins called cyclins. One of these, cyclin D1, in combination with its corresponding cyclin-dependent kinase (cdk), is essential for G(1)/S phase transition. Cyclin/cdk complexes are generally inhibited by cyclin-dependent kinase inhibitors(ckis), some of which are induced by wild-type p53. The aims of this study were: to investigate levels of cyclin D1 expression in transitional cell carcinoma (TCC) of the bladder; to correlate these results with data concerning the expression of p53, waf1, pRb and Ki67; and to determine whether cyclin D1 expression could predict clinical outcome. Paraffin-sections from 150 newly diagnosed bladder tumours (Ta/T1 = 97; T2-T4 = 53) were stained for cyclin D1 using immunohistochemistry and a cyclin D1 index assigned. These results were correlated with data relating to the expression of p53 and waf1 by the same tumours. A representative subset of 54 tumours (Ta/T1 = 28; T2-T4 = 26) was also stained for Ki67 and 55 were stained for pRb. The clinical course of each patient was recorded and multivariate analyses of risk factors for tumour recurrence, stage progression and overall survival were performed. Positive staining for cyclin D1 was found in 83% of tumours. The staining pattern varied between tumours with nuclear, cytoplasmic or a combination of the two evident in different tumours. 89% of Ta/T1 and 74% of T2-T4 tumours showed nuclear staining with or without cytoplasmic staining. The median value for cyclin D1 staining was significantly higher in Ta/T1 tumours (41%) compared with T2-T4 tumours (8%, P< 0.005) with 26% of muscle-invasive tumours demonstrating absent staining. In addition, the median value for cyclin D1 staining was significantly higher in G1/G2 tumours (43%) compared with G3 tumours (14%, P< 0.005). There was a significant positive correlation between expression of cyclin D1 and waf1 expression (P< 0.0001) as well as pRb expression but not between cyclin D1 expression and expression of p53. Ki67 expression was significantly associated with increasing tumour stage (P< 0.005) and histological grade (P< 0.05) but did not correlate with cyclin D1 expression. A cyclin D1 index > or = 8% was associated with significantly better survival in those patients with muscle-invasive disease (T2-T4). In addition, there was a significantly higher progression rate for those patients with Ta/T1 disease whose tumours demonstrated cytoplasmic cyclin D1 staining. These results indicate that cyclin D1 expression is significantly higher in low-stage, well differentiated bladder tumours and strongly correlates with waf1 expression. In a multivariate analysis, cyclin D1 expression is an independent prognostic indicator of survival in those patients with muscle-invasive disease.

Sugarcane yellow leaf virus: a novel member of the Luteoviridae that probably arose by inter-species recombination.

The 5895 nucleotide long single-stranded RNA genome of Sugarcane yellow leaf virus Florida isolate (SCYLV-F) includes six major ORFs. All but the first of these are homologous to genes of known function encoded by viruses of the three newly defined genera in the LUTEOVIRIDAE: ('luteovirids'), i.e. poleroviruses, luccccteoviruses and the enamoviruses. SCYLV-F ORFs 1 and 2 are most closely related to their polerovirus counterparts, whereas SCYLV-F ORFs 3 and 4 are most closely related to counterparts in luteovirus genomes, and SCYLV-F ORF5 is most closely related to the read-through protein gene of the only known enamovirus. These differences in affinity result from inter-species recombination. Two recombination sites in the genome of SCYLV-F map to the same genomic locations as previously described recombinations involving other luteovirids. A fourth type of luteovirid, Soybean dwarf virus, has already been described. Our analyses indicate that SCYLV-F represents a distinct fifth type.

Multi-step evolution of lung cancer.

In an attempt to improve the poor survival rates for lung cancer, therapeutic strategies require a deeper understanding of the biological events contributing to the formation and progression of the disease. In particular, the importance of studying the different stages of lung cancer including early pre-neoplasia is being recognised and studies examining genetic changes in pre-invasive and invasive lesions are being used to identify key events in tumorigenesis.

Risks associated with tattooing and body piercing.

The purpose of this article is to highlight the importance of including cosmetic body-piercing and tattooing in HIV prevention and education. Little information about risks associated with tattoos or having one's body pierced is evident in the health promotion and disease prevention literature, particularly among adolescents, ethnic groups, and incarcerated populations. It is incumbent that preventionists address behaviors such as tattooing and body piercing as possible vectors for HIV transmission in addition to typical concerns (homosexuality, i.v. drug use, condom use and safer sex practices). This article draws attention to the need for formation of regulatory policy issues related to body piercing and tattooing parlors. Currently, 26 percent of the states have regulatory authority over tattooing establishments, while only 4 states exercise such authority over body-piercing establishments. Implications for future research and policy initiatives are identified.

Prisoners' knowledge and attitude toward prophylactic treatment and therapy.

Identifying factors that hinder an inmate's compliance with Pneumocystis carinii pneumonia (PCP) prophylaxis therapy can be critical in preventing or decreasing the occurrence of PCP in this population. Anticipated factors include lack of knowledge about PCP and its proposed treatment, fear of the adverse effects of prophylaxis therapy, and lack of trust in the correctional facility medical team. Structured interviews were administered to HIV-positive male inmates chosen randomly (n = 104) at a medium- to maximum-security medical correctional facility located in the western portion of the United States. A basic "HIV 101 and Early Intervention" program encompassed the presentation of HIV facts and knowledge as well as safer sex practices. The results revealed that 95% of the respondents were knowledgeable about PCP and the side effects of their medications, and 56% of the respondents were afraid of the medications' side effects. Significant differences based on age were recorded for several specific knowledge questions, including the preventable nature of PCP.

Evidence that galactanase A from Pseudomonas fluorescens subspecies cellulosa is a retaining family 53 glycosyl hydrolase in which E161 and E270 are the catalytic residues.

A genomic library of Pseudomonas fluorescens subsp. cellulosa DNA was screened for galactanase-positive recombinants. The nine galactanase positive phage isolated contained the same galactanase gene designated galA. The deduced primary structure of the enzyme (galactanase A; GalA) encoded by galA had a Mr of 42 130 and exhibited significant sequence identity with a galactanase from Aspergillus aculeatus, placing GalA in glycosyl hydrolase family 53. The enzyme displayed properties typical of an endo-beta1, 4-galactanase and exhibited no activity against the other plant structural polysaccharides evaluated. Analysis of the stereochemical course of 2,4-dinitrophenyl-beta-galactobioside (2,4-DNPG2) hydrolysis by GalA indicated that the galactanase catalyzes the hydrolysis of glycosidic bonds by a double displacement general acid-base mechanism. Hydrophobic cluster analysis (HCA) suggested that family 53 enzymes are related to the GH-A clan of glycosyl hydrolases, which have an (alpha/beta)8 barrel structure. HCA also predicted that E161 and E270 were the acid-base and nucleophilic residues, respectively. Mutants of GalA in which E161 and E270 had been replaced with alanine residues were essentially inactive against galactan. Against 2,4-DNPG2, E161A exhibited a much lower Km and kcat than native GalA, while E270A was inactive against the substrate. Analysis of the pre-steady-state kinetics of 2,4-DNPG2 hydrolysis by E161A showed that there was an initial rapid release of 2,4-dinitrophenol (2,4-DNP), which then decayed to a slow steady-state rate of product formation. No pre-steady-state burst of 2,4-DNP release was observed with the wild-type enzyme. These data are consistent with the HCA prediction that E161 and E270 are the acid-base and nucleophilic catalytic residues of GalA, respectively.

Mannanase A from Pseudomonas fluorescens ssp. cellulosa is a retaining glycosyl hydrolase in which E212 and E320 are the putative catalytic residues.

Mannanase A (MANA) from Pseudomonas fluorescens, a member of glycosyl hydrolase family 26, was hyperexpressed in Escherichia coli and purified to homogeneity. Analysis of the stereochemical course of mannotetraose hydrolysis by purified MANA showed that the configuration of the anomeric carbon was retained on cleavage of the middle glycosidic bond. These data suggest that the mannanase hydrolyzes mannooligosaccharides by a double-displacement general acid-base mechanism. By hydrophobic cluster analysis (HCA), two glutamate and two aspartate residues were shown to be conserved in all of the glycosyl hydrolase family 26 enzymes analyzed. In addition, HCA suggested that family 26 was related to the GH-A clan (families 1, 2, 5, 10, 30, 35, 39, and 42) of (alpha/beta)8-barrel glycosyl hydrolases, which led to the prediction that E320 and E212 constitute the catalytic nucleophile and acid-base residues, respectively. To investigate the role of these amino acids, site-directed mutagenesis was used to replace the two aspartates with alanine and glutamate, while the two conserved glutamates were changed to alanine and aspartate. The mutant enzymes were purified and their biochemical properties were analyzed. The data showed that neither the D-->A nor the D-->E mutation resulted in a dramatic decrease in enzyme activity, suggesting that the two aspartate residues did not play a pivotal role in catalysis. In contrast, modification of either of the glutamate residues to alanine caused a dramatic decrease in kcat against carob galactomannan, azo-carob galactomannan, mannotetraose and 2,4-dinitrophenyl beta-mannobioside (2,4-DNPM). The E320A mutation did not alter the apparent K(m) (K(m)) of MANA against these substrates, while E212A resulted in a 27-fold decrease in K(m) against 2,4-DNPM. Pre-steady-state kinetics of 2,4-DNPM hydrolysis by E212A showed that there was a rapid burst of 2,4-dinitrophenol release. Circular dichroism and fluorescence spectroscopy indicated that there were no significant differences between the structures of the mutant and wild-type forms of MANA. These data are consistent with E212 and E320 constituting the catalytic acid-base and nucleophile residues of MANA, respectively.

A non-modular endo-beta-1,4-mannanase from Pseudomonas fluorescens subspecies cellulosa.

Pseudomonas fluorescens subsp. cellulosa when cultured in the presence of carob galactomannan degraded the polysaccharide. To isolate gene(s) from P. fluorescens subsp. cellulosa encoding endo-beta-1,4-mannanase (mannanase) activity, a genomic library of Pseudomonas DNA, constructed in lambda ZAPII, was screened for mannanase-expressing clones using the dye-labelled substrate, azo-carob galactomannan. The nucleotide sequence of the pseudomonad insert from a mannanase-positive clone revealed a single open reading frame of 1257 bp encoding a protein of M(r) 46,938. The deduced N-terminal sequence of the putative polypeptide conformed to a typical prokaryotic signal peptide. Truncated derivatives of the mannanase, lacking 54 and 16 residues from the N- and C-terminus respectively of the mature form of the enzyme, did not exhibit catalytic activity. Inspection of the primary structure of the mannanase did not reveal any obvious linker sequences or protein motifs characteristic of the non-catalytic domains located in other Pseudomonas plant cell wall hydrolases. These data indicate that the mannanase is non-modulator, comprising a single catalytic domain. Comparison of the mannanase sequence with those in the SWISSPROT database revealed greatest sequence homology with the mannanase from Bacillus sp. Thus the Pseudomonas enzyme belongs to glycosyl hydrolase Family 26, a family containing mannanases and endoglucanases. Analysis of the substrate specificity of the mannanase showed that the enzyme hydrolysed mannan and galactomannan, but displayed little activity towards other polysaccharides located in the plant cell wall. The enzyme had a pH optimum of approx. 7.0, was resistant to proteolysis and had an M(r) of 46,000 when expressed by Escherichia coli.

A novel family of cathepsin L-like (CTSLL) sequences on human chromosome 10q and related transcripts.

We have isolated a human genomic DNA cosmid clone while screening for the cathepsin L gene that, when sequenced, revealed close similarity with but significant differences from cDNA sequences that have been reported for cathepsin L (CTSL). The clone bears a novel sequence that shows 88% identity to the coding regions of the cathepsin L gene and a similar exon arrangement. We have called this sequence the "human cathepsin L-like gene 1" (CTSLL1). Translating putative exon sequences reveals a single premature stop codon; therefore no functional products are likely to arise from this gene. Fluorescence in situ hybridization (FISH) studies mapped the clone to chromosome 10q. Somatic cell hybrid mapping confirmed the location of CTSLL1 to human chromosome 10 distinct from the cathepsin L locus (CTSL) on chromosome 9. Furthermore, the FISH mapping studies show that a family of at least three related sequences exists on chromosome 10q, similar to the pattern of duplicated glutamate dehydrogenase (GLUD) gene loci reported on 10q. Using PCR and sequencing with genomic DNA samples, we have identified two additional novel related sequences (CTSLL2 and CTSLL3), and by PCR analysis of cDNA samples we have identified corresponding transcripts. Comparison of changes between our CTSLL1 sequence and the cathepsin L gene at mutation insensitive sites suggests that the two sequences arose from a duplication event 40-50 million years ago, and therefore at the time of divergence of early primates.

Teaching children about their internal bodies.

Children's knowledge about their internal bodies is related to their developmental age and the manner in which they are taught. Nurses need to consider appropriate methods, language, and timing for teaching basic anatomy and physiology to early school-age children to increase its effectiveness.