Genetic Diversity and Protective Efficacy of the RTS,S/AS01 Malaria Vaccine.

BACKGROUND: The RTS,S/AS01 vaccine targets the circumsporozoite protein of Plasmodium falciparum and has partial protective efficacy against clinical and severe malaria disease in infants and children. We investigated whether the vaccine efficacy was specific to certain parasite genotypes at the circumsporozoite protein locus. METHODS: We used polymerase chain reaction-based next-generation sequencing of DNA extracted from samples from 4985 participants to survey circumsporozoite protein polymorphisms. We evaluated the effect that polymorphic positions and haplotypic regions within the circumsporozoite protein had on vaccine efficacy against first episodes of clinical malaria within 1 year after vaccination. RESULTS: In the per-protocol group of 4577 RTS,S/AS01-vaccinated participants and 2335 control-vaccinated participants who were 5 to 17 months of age, the 1-year cumulative vaccine efficacy was 50.3% (95% confidence interval [CI], 34.6 to 62.3) against clinical malaria in which parasites matched the vaccine in the entire circumsporozoite protein C-terminal (139 infections), as compared with 33.4% (95% CI, 29.3 to 37.2) against mismatched malaria (1951 infections) (P=0.04 for differential vaccine efficacy). The vaccine efficacy based on the hazard ratio was 62.7% (95% CI, 51.6 to 71.3) against matched infections versus 54.2% (95% CI, 49.9 to 58.1) against mismatched infections (P=0.06). In the group of infants 6 to 12 weeks of age, there was no evidence of differential allele-specific vaccine efficacy. CONCLUSIONS: These results suggest that among children 5 to 17 months of age, the RTS,S vaccine has greater activity against malaria parasites with the matched circumsporozoite protein allele than against mismatched malaria. The overall vaccine efficacy in this age category will depend on the proportion of matched alleles in the local parasite population; in this trial, less than 10% of parasites had matched alleles. (Funded by the National Institutes of Health and others.).

Interest in multipurpose prevention of HIV and pregnancy: perspectives of women, men, health professionals and community stakeholders in two vaginal gel studies in southern Africa.

OBJECTIVE: The pipeline of vaginal microbicides for HIV prevention has expanded to include products for multipurpose prevention, but the interests of potential users and those advising on use have not been sufficiently investigated. Rather, assumptions about interest in multipurpose prevention technologies (MPTs) are inferred from what is known about acceptability and use of microbicides or contraceptives. DESIGN AND SETTING: This paper presents data on concerns and preferences for multipurpose prevention of HIV and pregnancy. Data were collected in two microbicide gel studies in Malawi and Zimbabwe. Participants were women using candidate vaginal products, their male partners, health professionals and community stakeholders. METHODS: An individual interview was conducted with participants. Interviews were audio-recorded, transcribed, coded for content and analysed for key themes. RESULTS: Participants indicated strong interest in a vaginal HIV prevention product that could also prevent pregnancy. Reasons for this interest were convenience, problems with adverse effects with current contraceptive methods, concerns about long-term effects of contraceptives, and concerns about the health burdens of HIV infection during pregnancy. The main disadvantage of an MPT was recognition that while interest in preventing HIV is constant, contraceptive needs change over time. CONCLUSION: The study population indicated support for an MPT to prevent HIV and pregnancy. This support may be further strengthened if the product is also available for prevention of only HIV. Women and men will be more willing to use an MPT if they can be reassured that its use will have no long-term effect on fertility.

Acceptance of repeat population-based voluntary counselling and testing for HIV in rural Malawi.

OBJECTIVE: To examine the acceptance of repeat population-based voluntary counselling and testing (VCT) for HIV in rural Malawi. METHODS: Behavioural and biomarker data were collected in 2004 and 2006 from approximately 3000 adult respondents. In 2004, oral swab specimens were collected and analysed using ELISA and confirmatory Western blot tests, while finger-prick rapid testing was done in 2006. We used cross-tabulations with chi(2) tests and significance tests of proportions to determine the statistical significance of differences in acceptance of VCT by year, individual characteristics and HIV risk. RESULTS: First, over 90% of respondents in each round accepted the HIV test, despite variations in testing protocols. Second, the percentage of individuals who obtained their test results significantly increased from 67% in 2004, when the results were provided in randomly selected locations several weeks after the specimens were collected, to 98% in 2006 when they were made available immediately within the home. Third, whereas there were significant variations in the sociodemographic and behavioural profiles of those who were successfully contacted for a second HIV test, this was not the case for those who accepted repeat VCT. This suggests that variations in the success of repeat testing might come from contacting the individuals rather than from accepting the test or knowing the results. CONCLUSIONS: Repeat HIV testing at home by trained healthcare workers from outside the local area, and with either saliva or blood, is almost universally acceptable in rural Malawi and, thus, likely to be acceptable in similar contexts.

Genotypic analysis of the protease and reverse transcriptase of HIV type 1 subtype C isolates from antiretroviral drug-naive adults in Malawi.

The protease (PR) and reverse transcriptase (RT) regions of HIV-1 isolates from 21 antiretroviral (ARV)-naive Malawian adults were sequenced and analyzed to determine the prevalence of drug resistance-associated mutations in this population. Phylogenetic analysis confirmed that all isolates grouped with HIV-1 subtype C, the predominant subtype in Malawi. No major mutations associated with resistance to PR inhibitors (PIs), nucleoside RT inhibitors (NRTIs), or nonnucleoside RT inhibitors (NNRTIs) were found. In contrast, accessory mutations were found in the protease region at positions 10, 20, 36, 63, 77, and 93, and in the RT region at positions 118, 211, and 214. Further studies will be needed to determine the clinical impact of these polymorphisms on viral susceptibility to existing antiretroviral drugs.

A dominant negative CREB (cAMP response element-binding protein) isoform inhibits thyrocyte growth, thyroid-specific gene expression, differentiation, and function.

cAMP mediates the effects of TSH by regulating thyroid follicular cell proliferation, differentiation, and function. To assess the functional importance of the cAMP response element binding protein (CREB) in thyroid follicular cell regulation in vivo, we targeted the expression of a dominant negative (DN) CREB isoform to the thyroid glands of transgenic mice using a tissue-specific promoter. Transgenic mice exhibited severe growth retardation and primary hypothyroidism. Serum levels of TSH were elevated 8-fold above normal levels, and T4 and T3 levels were low. Histologically, the mutant thyroid glands were characterized by poorly developed follicles that were heterogeneous in size with diminished colloid. Ciliated thyroid epithelial cells were observed in the transgenic thyroid glands, suggesting a failure of follicular cell differentiation. Consistent with this hypothesis, the DN CREB transgene inhibited the expression of an array of genes including thyroglobulin, thyroperoxidase, and the TSH receptor in semiquantitative RT-PCR experiments. Altered expression of the thyroid transcription factors Pax-8, TTF-1, and TTF-2 was also observed. These results demonstrate a critical role for CREB in thyroid growth, differentiation, and function in vivo.

Blockage of the rete testis and efferent ductules by ectopic Sertoli and Leydig cells causes infertility in Dax1-deficient male mice.

DAX-1, an X-linked member of the orphan nuclear receptor superfamily of transcription factors, plays a key role in sex determination and gonadal differentiation. Dax1-deficient male mice are infertile and have small testes despite normal serum levels of T and gonadotropins. Examination of Dax1-deficient testes reveals dilated seminiferous tubules and abnormal parameters of sperm fertilizing capability consistent with a possible obstruction in the testis. To test this hypothesis, we performed a comprehensive evaluation of the male reproductive tract in Dax1-deficient mice. Light and electron microscopic examination revealed the rete testis is blocked by aberrantly located Sertoli cells, creating a tailback of necrosing sperm in the testis. Sertoli cells also obstruct the proximal and middle efferent ductules, and this is accompanied by an overgrowth of the efferent duct epithelium. Seminiferous tubules close to the rete testis contain ectopic Leydig cells, distinct from the hyperplastic Leydig cells present in the interstitial space. The peritubular tissue surrounding these tubules is frequently abnormal, containing relatively undifferentiated myoid cells and no basement membrane between the myoid cells and Sertoli cells. A third of aged (>1-yr-old) Dax1-deficient male mice develop sex cord-stromal tumors, derived from cells of the Sertoli/granulosa cell or Leydig cell lineages. Combined, these observations reveal abnormal differentiation and proliferation of Leydig cells and Sertoli cells in Dax1-deficient male mice, leading to obstruction of the rete testis and infertility.

Sertoli cell-specific rescue of fertility, but not testicular pathology, in Dax1 (Ahch)-deficient male mice.

DAX1 is an orphan member of the nuclear hormone receptor superfamily of transcription factors. Our recent characterization of Dax1 (Ahch)-deficient male mice revealed a primary testicular defect resulting in hypogonadism and sterility. The progressive degeneration of the germinal epithelium, independent of abnormal gonadotropin and testosterone production, suggested an intrinsic loss of Dax1 function in the Sertoli cells. To test this hypothesis, we assessed the effect of Sertoli cell-specific expression of a human DAX1 (AHC) transgene driven using the promoter of the Müllerian inhibiting substance (MIS) gene. The MIS-DAX1 transgene partially rescued the mutant phenotype of the Dax1-deficient male mice. Although testicular morphology remained abnormal, fertility was restored to levels matching that of wild-type littermates. Examination of several markers of sperm fertilizing capability revealed significant improvements in MIS-DAX1-rescued mice. Epididymal sperm count and sperm motility were greater in 12-week-old rescued mice than in age-matched Dax1-deficient mice. The ability of sperm to undergo an immediate acrosome reaction was impaired in Dax1-deficient animals, and sperm from Dax1-deficient mice fertilized only 8.2 +/- 6.8% of eggs in vitro, significantly less than rescue (67.8 +/- 19.1%) and wild-type (88.9 +/- 3.9%) sperm. These results indicate that Dax1 expression in Sertoli cells is adequate to overcome crucial thresholds related to sperm production and function. However, the failure to completely rescue the testicular pathology of Dax1-deficient mice suggests that Dax1 expression in other somatic cells is essential for normal testicular development.

Adenovirus-mediated targeted expression of toxic genes to adrenocorticotropin-producing pituitary tumors using the proopiomelanocortin promoter.

Management of Cushing's disease remains challenging, despite advances in its diagnosis and treatment. Here, we describe a strategy for targeting the expression of toxic genes to ACTH-producing tumor cells using adenoviral vectors. The POMC promoter was used to express either a marker gene (beta-galactosidase) or a toxic gene [herpes simplex virus thymidine kinase (TK)]. In ACTH-producing AtT20 cells, infection with recombinant adenoviruses containing the POMC promoter (AdPOMCGal; AdPOMCTK) led to high-level gene expression. Stereotactic injection of AdPOMCGal into the rat pituitary resulted in localized expression of the beta-galactosidase transgene in corticotrope cells. Cytotoxicity studies were performed using the TK-containing vectors and treatment with ganciclovir. AdPOMCTK caused greater than 95% cytotoxicity of AtT20 cells at a viral dose (multiplicity of infection, 5 plaque-forming units/cell) that induced minimal toxicity using control viruses. No cellular toxicity was seen using a nonpituitary cell line (T47D breast tumor cells). AtT20 cells transplanted into nude mice induced features of Cushing's syndrome and were used as an in vivo model of ACTH-producing tumors. Injection of the AdPOMCTK virus caused significant regression of the transplanted AtT20 tumors. These studies suggest that the POMC promoter may provide a useful gene therapy strategy for the adjunctive treatment of pituitary tumors causing ACTH-dependent Cushing's syndrome.

Vagal afferent innervation of smooth muscle in the stomach and duodenum of the mouse: morphology and topography.

Intraganglionic laminar endings (IGLEs) and intramuscular arrays (IMAs), the two putative mechanoreceptors that the vagus nerve supplies to the gastrointestinal smooth muscle, have been characterized almost exclusively in the rat. To provide normative inventories of these afferents for the mouse, the authors examined the endings in the stomach and small intestine of three strains used as backgrounds for gene manipulations (i.e., C57, 129/SvJ, and WBB6). Animals received nodose ganglion injections of wheat germ agglutinin-horseradish peroxidase or dextran-tetramethylrhodamine conjugated to biotin. The horseradish peroxidase tissue was processed with tetramethylbenzidine and was used to map the distributions and densities of the two endings; the dextran material was counterstained with c-Kit immunohistochemistry to assess interactions between intramuscular arrays and interstitial cells of Cajal. IGLEs and IMAs constituted the vagal innervation of mouse gastric and duodenal smooth muscle. IGLE morphology and distributions, with peak densities in the corpus-antrum, were similar in the three strains of mice and comparable to those observed in rats. IMAs varied in complexity from region to region but tended to be simpler (fewer telodendria) in mice than in rats. IMAs were most concentrated in the forestomach and sphincters in mice, as in rats, but the topographic distributions of the endings varied both between strains of mice (subtly) and between species (more dramatically). IMAs appeared to make appositions with both interstitial cells and smooth muscle fibers. This survey should make it practical to assay the effects of genetic (e.g., knockout) and experimental (e.g., regeneration) manipulations affecting visceral afferents and their target tissues.

Correlation between biomarkers of omega-3 fatty acid consumption and questionnaire data in African American and Caucasian United States males with and without prostatic carcinoma.

Results from animal studies suggest that omega-3 fatty acids from marine sources are protective against cancer. To determine whether adipose tissue and erythrocyte membrane fatty acid composition could serve as biomarkers of essential fatty acid consumption in subjects with prostate cancer, we compared fish consumption, which was estimated using a food frequency survey, to the omega-3 fatty acid content of adipose tissue and erythrocyte membranes. The study was conducted using 127 men who had undergone a prostate biopsy. All subjects were recruited from a university hospital urology clinic. African Americans comprised 23% of the subjects, and 70% were diagnosed with prostate cancer. We found a correlation of 0.44 with 95% confidence intervals (CIs) = 0.29-0.57 between reported fish consumption and the omega-3 fatty acid eicosapentaenoic acid composition in erythrocyte membranes and 0.38 with 95% CI = 0.21-0.53 when the dietary survey was compared to eicosapentaenoic acid in adipose tissue. The survey/biomarker correlations in cases were not significantly different from the correlations in controls. The study had 90% power to detect a 0.35 difference between correlations. These results suggest that the presence of prostate cancer does not affect the adipose tissue or erythrocyte membrane biomarkers of fatty acid consumption, and that erythrocyte membranes are as useful as biomarkers as is adipose tissue. Our findings corroborate previous studies that found that tissue biomarkers can reflect past fatty acid consumption and support the use of biomarkers in case-control studies using cancer patients.

Biomarkers of essential fatty acid consumption and risk of prostatic carcinoma.

Animal studies have suggested that omega-6 fatty acids found in vegetable oils may promote prostate cancer. Our goal was to use erythrocyte membrane and adipose tissue fatty acid composition as biomarkers to investigate whether essential fatty acids modulated prostate cancer risk. An outpatient clinic-based study of 89 cases and 38 controls was conducted in North Carolina between July 1989 and December 1991. Cases were recruited from a university-based urology outpatient clinic. Eligible cases were more than 45 years of age and had histological confirmation of a prostate cancer diagnosis within 1 year of entry into the study. Controls were histologically confirmed free of prostate cancer. Erythrocyte membranes from venous blood and adipose tissue fatty acids from s.c. fat samples were analyzed in batches using capillary gas chromatography. Unconditional logistic regression analysis was used to calculate odds ratios for the association of each fatty acid with prostate cancer while controlling for potential confounders. Linoleic acid consumption was positively associated with prostate cancer risk. The odds ratios comparing the first and fourth quartiles of linoleic acid consumption were 3.54 (95% confidence interval, 1.0-12.53) with P trend < 0.04 for erythrocyte membranes, and 2.47 (95% confidence interval, 0.66-9.26) with P trend < 0.08 for adipose tissue. These data suggest that linoleic acid consumption may increase prostate cancer risk, which is consistent with results from animal experiments. Linoleic acid is found in vegetable oils used in cooking and in cereals, snack foods, and baked goods. Our data failed to demonstrate consistently a protective effect of marine omega-3 fatty acids on prostate cancer.

Driver's license and voter registration lists as population-based sampling frames for rural African Americans.

PURPOSE: To compare coverage of a state driver's license list and county voter registration lists as frames for sampling rural African Americans. METHODS: Name, birth date, sex, and county were extracted from records for all 18--59 year-old African Americans residing in eight rural North Carolina counties and listed in the North Carolina Driver license file [obtained as a SAS dataset from the University of North Carolina (UNC) Highway Safety Research Center] and in machine-readable text files of registered voters (obtained from county boards of elections). Files were edited, merged, and matched by last name and date of birth to produce three files for each five-year age group, sex, and county: 1) persons listed only in the driver's file; 2) persons listed only in the voter's file; and 3) persons listed in both. RESULTS: The median percentages of unique persons found only on the driver's list, only on the voter's list, and on both lists were 54%, 17%, and 30% men, and 35%, 25%, and 41% women, respectively. CONCLUSIONS: The driver's list provided better coverage than did voter registration lists. Federal legislation that prohibits states from releasing driver's license lists for use in surveys removes a valuable resource for population-based research.

Habituation of oral responding in adult rats.

The effects of repeated oral stimulation on ingestive responding were investigated in adult rats. A series of brief intraoral infusions of flavored diet was delivered to female rats once every minute through an oral cannula. When the flavor of the infused diet remained constant, significant decreases in mouthing behavior were observed by the end of testing, whereas switching the flavor of the diet during testing resulted in enhanced responding and infusions delivered through gastric cannulas produced minimal effects. Patterns of oral responding were also similar in food-restricted rats. These patterns of responding suggest that adult rats habituate to oral stimulation. Finally, oral habituation led to decreased ingestion, whereas gastric infusions had minimal effects. Thus, oral habituation may represent a mechanism influencing intake in rats at all ages.

Gastrointestinal projection maps of the vagus nerve are specified permanently in the perinatal period.

The vagal innervation of the proximal gastrointestinal (GI) tract is lateralized. To determine whether this pattern is specified as early as the perinatal period, neonatal rat pups were given unilateral cervical vagotomies. Separate groups received (1) transections below the left nodose ganglion, (2) left cervical resections that included removal of the nodose ganglion, or (3) sham surgeries. At 4 months of age, each animal's vagal afferent projections from the unoperated side were mapped by injecting the nodose with WGA-HRP, preparing the stomach as wholemounts, and processing the tissue with tetramethyl benzidine. The two types of vagal afferent endings in GI smooth muscle, namely intraganglionic laminar endings and intramuscular arrays, were surveyed separately, and their regional distributions were mapped. Changes in the nucleus of the solitary tract (NST) and dorsal motor nucleus of the vagus (DMNX) were assessed with cell counts and area measurements. Neonatal loss of the vagus innervating one side of the GI tract, with or without ganglionectomy, did not cause the unoperated vagus to sprout to the denervated side. In addition, removal of the projections to the one side of the target organ did not produce a reorganization of the projection maps of the unoperated vagus within its normal or ipsilateral wall of the GI tract. Although the regional patterns of the unoperated ipsilateral vagus were not affected, the packing densities of both types of afferents supplied by this trunk were moderately reduced. The DMNX of the vagotomized side displayed extensive (approximately 83%) neuronal loss; the DMNX on the unoperated side as well as the NST on both sides exhibited limited (approximately 20--25%) losses. The lack of a peripheral projection field reorganization -- except for a moderate down-regulation -- after complete unilateral denervation suggests that both the laterality and the afferent terminal phenotypes (or target tissues) of the vagus in the proximal GI tract are specified by postnatal day one in the rat. The present results, taken together with other observations, also suggest that three different combinations of signals orchestrate the commitments of vagal afferents respectively to (1) the side of the organ, (2) the region within the organ wall, and (3) the accessory and innervated tissues that complex with the fully differentiated ending.

C-Kit mutant mice have a selective loss of vagal intramuscular mechanoreceptors in the forestomach.

Intramuscular arrays are one of two major classes of vagal afferent mechanoreceptors that innervate the smooth muscle wall of the proximal gastrointestinal tract. They consist of rectilinear telodendria that distribute in the muscle sheets, parallel to the long axes of muscle fibers. Intramuscular arrays appear to make direct contact with the muscle fibers, but they also course on, and form appositions with, intramuscular interstitial cells of Cajal. These complexes formed by intramuscular arrays and intramuscular interstitial cells of Cajal suggest that intramuscular arrays might require either structural or trophic support of the interstitial cells of Cajal for normal differentiation and/or maintenance. To evaluate this hypothesis, we have examined the morphology and distribution of vagal afferent endings in the c-Kit mutant mouse that lacks intramuscular interstitial cells of Cajal. Vagal afferents were labeled by nodose ganglion injection of either wheat germ agglutinin-horseradish peroxidase conjugate or a tagged dextran, and the labeled afferent terminals in the stomach were mapped using a standardized quantitative sampling scheme. Intramuscular arrays were dramatically reduced (in circular muscle by 63%; in longitudinal muscle by 78%) in the c-Kit mutant mice relative to their wild-type littermates. Additionally, a substantial number of the surviving axons and terminals in the mutant stomachs were morphologically aberrant. Moreover, the loss of intramuscular arrays in mutants appeared to be selective: the structure, distribution and density of intraganglionic laminar endings, i.e., the other vagal mechanoreceptors in smooth muscle, were not significantly altered. Finally, the conspicuous decrease in intramuscular array density in mutants was associated with a non-significant trend toward loss of nodose ganglion neurons. Collectively these findings suggest that interstitial cells are required for the normal development or maintenance of vagal intramuscular arrays. Therefore, the c-Kit mutant mouse will be valuable for determining the role(s) of interstitial cells in intramuscular array development as well as for providing an animal model with the intramuscular array class of vagal afferents selectively ablated.

Tomography of the petrous bone in keratosis obturans.

Three cases of keratosis obturans, which were studied by tomography of the petrous temporal bone, are described. The widening of the deep bony part of the external auditory canal by pressure erosion of an impacted benign slowly-growing mass is the key radiological finding. Erosion was so severe as to involve the facial nerve canal in one and the temporo-madibular joint in two cases. The theories of causation are reviewed. Tomography using elliptical tube movement was valuable in diagnosis, pre- and post-operative management.

A study of deafness in West Africa.

Some 800 profoundly deaf children from 4 Yoruba states in Nigeria were assessed and examined in an attempt to identify the causes of deafness in this part of West Africa. It was found that measles, meningitis and rubella were the three major preventible diseases causing deafness. Genetic abnormalities, in contradistinction to other tropical countries, do not seem to be very important and chronic ear infections show different manifestations in this area to the situation reported in East Africa. The overwhelming problems that face the deaf and any possible extension of services for the deaf are detailed. In view of these it seems that the most cost-effective way of helping the deaf in Nigeria is by immunizing the population against those infectious diseases that cause the disability. This policy would accord with the recent initiatives by the United Nations: the International Year of the Child (I.Y.C.) and the International Year of the Disabled Person (I.Y.D.P.) as well as The Expanded Programme of Immunization (E.P.I.) of the World Health Organization.

Chemodectomas: a review of 17 cases.

Seventeen cases of chemodectomas seen in the University College Hospital, Ibadan, are presented. Eight of them (48%) are carotid body tumors, while six (36%) are glomus jugulare. The rest are subglottic, abdominal and multicentric. The sex distribution for the two major types is equal for males and females; but they all seem to appear in younger age groups. Operative mortality has been exclusive to carotid body tumors (50% for this group), while bleeding is the leading postoperative complication in all the cases. The embryology, pathophysiology and clinical presentation of chemodectomas are discussed and it is suggested that, in the management of this condition, the advantages of operation should be weighed against morbidity and mortality risks.

Otolaryngological complications of measles in West Africa.

In Nigeria as in West Africa generally measles is a serious condition with a high morbidity and mortality rate. Complications involve every section of Otolaryngology. Some of these are amenable to treatment by the otolaryngologist alone, while others need the skill of colleagues in other specialties. The irreversible neuro-audiological complications cause greatest concern to physician and parent and will continue to do so until the variety of available therapeutic, preventive and rehabilitative services are increased and improved.

Nasopharyngeal angiofibroma.

Three cases of nasopharyngeal angiofibroma have been described along with the histopathological findings, treatment and a brief review of the literature on the subject. It is of interest that one of these cases, histologically proven, was a female and this in spite of some rigidly held views about the strict sex linkage of the lesion. The tumour is uncommon but it is possible that unfamiliarity with the histopathology, as well as the likelihood of cessation of activity at any stage, may in part be responsible for its low recorded prevalence. The progress of each patient after four, two and one year respectively gives cause for some optimism, but the long-term prognosis must remain guarded.