Multiplex PCR and Microarray for Detection of Swine Respiratory Pathogens.

Porcine respiratory disease complex (PRDC) is one of the most important health concerns for pig producers and can involve multiple viral and bacterial pathogens. No simple, single-reaction diagnostic test currently exists for the simultaneous detection of major pathogens commonly associated with PRDC. Furthermore, the detection of most of the bacterial pathogens implicated in PRDC currently requires time-consuming culture-based methods that can take several days to obtain results. In this study, a novel prototype automated microarray that integrates and automates all steps of post-PCR microarray processing for the simultaneous detection and typing of eight bacteria and viruses commonly associated with PRDC is described along with associated multiplex reverse transcriptase PCR. The user-friendly assay detected and differentiated between four viruses [porcine reproductive and respiratory syndrome virus (PRRSV), influenza A virus, porcine circovirus type 2, porcine respiratory corona virus], four bacteria (Mycoplasma hyopneumoniae, Pasteurella multocida, Salmonella enterica serovar Choleraesuis, Streptococcus suis), and further differentiated between type 1 and type 2 PRRSV as well as toxigenic and non-toxigenic P. multocida. The assay accurately identified and typed a panel of 34 strains representing the eight targeted pathogens and was negative when tested with 34 relevant and/or closely related non-target bacterial and viral species. All targets were also identified singly or in combination in a panel of clinical lung samples and/or experimentally inoculated biological material.

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.).

Laminar and spatial localization of the islands of Calleja in mice.

The islands of Calleja (IC) are dense clusters of cells localized within the ventral striatum. The IC have been described as variable in both number and localization from animal-to-animal, however, a quantitative investigation of this variability is unavailable. Further, it is presently unknown whether the IC occupy select areas of the olfactory tubercle (OT), the ventral striatum structure which possesses the IC in mice. To address these questions, we examined the IC of adult C57bl/6 mice. As previously noted, we found substantial inter-hemispheric and inter-mouse variations in the total number of IC. While the IC were observed in all three cell layers of the OT, the bulk of IC occupied layer iii. The span of the IC along the anterior-posterior and medial-lateral axes of the OT was variant. Further, localizations of the IC within the OT also differed across animals. Notably, the probability of observing an IC in the medial OT was greater than that of observing one in the lateral. These data provide a fundamental characterization of both differences and similarities regarding the IC in mice and will be informative for future in vivo studies seeking to perturb and possibly record from the IC. Further, we predict that inter-animal diversity in the IC may be a mechanism for inter-animal differences in behavior, especially reward-related and motivational behaviors.

Unsweetened natural cocoa has anti-asthmatic potential.

Unsweetened natural cocoa powder is enriched with nutraceutical abundance of anti-asthmatic compounds theobromine and theophylline. Cocoa powder, which is prepared after removal of the cocoa butter, contains about 1.9% theobromine and 0.21% caffeine. Anecdotal reports indicate that regular consumption of unsweetened natural cocoa powder (UNCP), a common practice in Ghana, West Africa, has the potential to reduce the tendency of asthmatic episodes. In the present paper we studied the effect of regular ingestion of aqueous extract of UNCP on hematological and histopathological changes that occur in ovalbumin (OVA)-sensitized guinea pigs. OVA-sensitized guinea pigs were challenged with aerosolized OVA 1 hour after ingestion of 300 mg/kg (low dose) or 600 mg/kg (high dose) of UNCP for 35 consecutive days. Histopathological and haematological changes in the OVA-sensitized guinea pigs were evaluated. Both negative and positive controls with distilled water and prednisolone, respectively, were used. OVA-sensitized guinea pigs demonstrated concentration-independent reduction in immune response to aerosolized OVA. There were no histo-architectural changes in the bronchiolar smooth muscles of the treated groups. Unsweetened natural cocoa powder has potential anti-asthmatic properties when administered orally at the doses tested.

Prostate-specific targeting of the aqueous root extract of Croton membranaceus in experimental animals.

Croton membranaceus Müll.Arg. (Euphorbiaceae) is used for benign prostate hyperplasia (BPH) treatment. The study aimed at investigating organs that the aqueous root extracts of C. membranaceus (CMARE) target, which is absent in literature. Twenty-four male Sprague-Dawley rats (100-140 g) were randomly divided into 4 groups. Group 1, the control group received distilled water. Groups 2, 3 and 4 received 30, 150 and 300 mg kg(-1) b.wt CMARE respectively (oral gavage). Rats fed 90 days the standard chow diet ad libitum. Upon sacrifice, major organs were histologically examined and serum prostate-specific antigen (PSA) biochemically determined. Only the prostate was abnormal. Histologically, H&E staining revealed thickness and infoldings of the epithelial cells shrinking with increasing dose. The 30 mg kg(-1) group showed low columnar or flattened epithelium cells, whereas the columnar epithelium infoldings of the 150 mg kg(-1) b.wt and 300 mg kg(-1) b.wt groups were virtually nonexistent. The acini of the control, 30 mg kg(-1) b.wt group and the 150 mg kg(-1) b.wt groups showed clear pinkish secretion. However, secretion of the high-dose group appeared light pink in colour and the stroma cells appeared much darker than all the treated and control group. C. membranaceus targets the prostate with significant PSA reduction (P < 0.01).

Genotoxicity, cytotoxicity and toxicological evaluation of whole plant extracts of the medicinal plant Phyllanthus niruri (Phyllanthaceae).

Phyllanthus niruri is a medicinal plant (commonly known as stone breaker) found in the tropics and other parts of the world. It is known for its capacity to block the formation of calcium oxalate crystals and kidney stone formation in urolithiasis. This plant has been used to treat hyperglycemia, hypertension, pain, and mild cases of malaria. We examined the geno-, cyto- and overall toxicity of P. niruri whole plant ethanolic extract. The extract was administered as a single dose of 30 or 300 mg/kg to laboratory rats by gavage, accompanied by negative (0.9% saline) and positive (10 mg/mL N-ethyl-N-nitrosourea) controls that were injected intramuscularly 48 h after extract administration. The ratio of polychromatic (PCE)/normochromatic erythrocytes (NCE) from femur bone marrow was scored for genotoxicity. Cytotoxicity was determined using descending concentrations (0.2-0.0125 g/mL) of the extract incubated with peripheral blood mononuclear cells. Lactate dehydrogenase release from damaged cells was determined and the CC(50) calculated. Subchronic administration of the extract at 30 or 300 mg/kg was done for 90 days to determine general toxicity. PCE:NCE (%) for the extract and negative control was 63, compared to 168 (positive control). The CC(50) was 26.3 mg/mL and hepato-renal toxicity after subchronic extract administration was nil. We conclude that ethanol extract of P. niruri is not cytotoxic or genotoxic, and is generally non-toxic on subchronic administration.

Expression of Plasmodium falciparum 3D7 STEVOR proteins for evaluation of antibody responses following malaria infections in naïve infants.

Clinical immunity to Plasmodium falciparum malaria develops after repeated exposure to the parasite. At least 2 P. falciparum variant antigens encoded by multicopy gene families (var and rif) are targets of this adaptive antibody-mediated immunity. A third multigene family of variant antigens comprises the stevor genes. Here, 4 different stevor sequences were selected for cloning and expression in Escherichia coli and His6-tagged fusion proteins were used for assessing the development of immunity. In a cross-sectional analysis of clinically immune adults living in a malaria endemic area in Ghana, high levels of anti-STEVOR IgG antibody titres were determined in ELISA. A cross-sectional study of 90 nine-month-old Ghanaian infants using 1 recombinant STEVOR showed that the antibody responses correlated positively with the number of parasitaemia episodes. In a longitudinal investigation of 17 immunologically naïve 9-month-old infants, 3 different patterns of anti-STEVOR antibody responses could be distinguished (high, transient and low). Children with high anti-STEVOR-antibody levels exhibited an elevated risk for developing parasitaemia episodes. Overall, a protective effect could not be attributed to antibodies against the STEVOR proteins chosen for the study presented here.

Evidence of increased diversity of methanogenic archaea with plant extract supplementation.

This study evaluated the effects of selected essential oils on archaeal communities using the ovine rumen model. Forty weaned Canadian Arcott ewes, fed with barley-based diet, were allotted to one of three essential oil supplementation treatments or a control (10 ewes per treatment) for 13 weeks. The treatments were cinnamaldehyde, garlic oil, juniper berry oil, and a control with no additive. Rumen content was sampled after slaughter and grouped by treatment by combining subsamples from each animal. DNA was extracted from the pooled samples and analyzed for methanogenic archaea using quantitative polymerase chain reaction, denaturing gradient gel electrophoresis, cloning, and sequencing. Our results suggest that the total copy number of archaeal 16S rRNA was not significantly affected by the treatments. The phylogenetic analysis indicated a trend toward an increased diversity of methanogenic archaea related to Methanosphaera stadtmanae, Methanobrevibacter smithii, and some uncultured groups with cinnamaldehyde, garlic, and juniper berry oil supplementation. The trends in the diversity of methanogenic archaea observed with the essential oil supplementation may have resulted from changes in associated protozoal species. Supplementation of ruminant diets with essential oils may alter the diversity of rumen methanogens without affecting the methanogenic capacity of the rumen.

Effect of insertional mutations in the pueA and pueB genes encoding two polyurethanases in Pseudomonas chlororaphis contained within a gene cluster.

AIMS: To better understand the role of PueA and PueB from Pseudomonas chlororaphis in polyurethane degradation, the present study was conducted to create insertional mutants in their respective genes. METHODS AND RESULTS: Growth kinetic studies showed that the pueA knockout mutant had a greater effect than the pueB knockout mutant. The pueA mutant had an 80% decrease in cell density from that of the wild type, while the pueB mutant had an 18% decrease in cell density. Polyurethane utilization followed Michaelis-Menten kinetics. The pueA and pueB mutants exhibited a 17% and 10% decrease respectively in growth rate using polyurethane when compared with the wild type. CONCLUSIONS: In this present study, pueA and pueB, are shown to be part of an ABC transporter gene cluster that consists of seven open reading frames. Mutational analysis results suggest that PueA may play a more major role in polyurethane degradation than PueB based on cell density and growth rates. SIGNIFICANCE AND IMPACT OF THE STUDY: The results from this study provide a starting point for the eventual enhancement and bioremediation of polyurethane waste. Understanding the role of polyurethane-degrading enzymes is useful for the creation of strains for this purpose.

Improving access to early treatment of malaria: a trial with primary school teachers as care providers.

The feasibility of improving access to early case detection and prompt and adequate management of acute episodes of malaria using school teachers was explored through an intervention trial in Ghana. Of all the 'fevers' diagnosed as presumptive malaria by the trained teachers, 93% met the case definition. However, a lower proportion (75%) of such correctly diagnosed cases were subsequently treated according to the treatment protocol provided. In a scaled up study, pre-packaging of the antimalarial drug improved the rate of adequate treatment to 97% of cases correctly diagnosed as presumptive malaria. Pre-packaging of chloroquine ensured a high level of user compliance (96.6%), even in the face of diminished supervision of the teachers. It is feasible for the health and education sectors to work in partnership to improve access to early case detection and adequate management of acute episodes of malaria.

Effect of vitamin E, low dose irradiation, and display time on the quality of pork.

A 4×2×3 factorial experiment was designed to investigate the effect of supplemental vitamin E (0, 100, 200, and 300 mg/kg feed), irradiation and days in display on quality characteristics of aerobically packaged ground pork and vacuum packaged loin chops. Color, thiobarbituric acid reactive substances (TBARS), olfactory and sensory characteristics were evaluated. Samples held in display for 3 d were used for sensory evaluation. In the ground pork, irradiation (1.9 kGy) increased "wetdog" flavor, increased a(*) (redness) and decreased L(*) (lightness) and b(*) (yellowness) values but had no effect on TBARS. However, as display time (0, 4, and 8 days) increased; the differences in a(*) values diminished and putrefying and fishy odors were higher in non-irradiated samples than irradiated. Supplemented vitamin E had no effect on TBARS, odor, and color measures; but increased the juiciness of ground pork regardless of irradiation. Similarly, in the loin chops, irradiation (1.5 kGy) increased a(*) value and "wetdog" flavor; but decreased b(*) value regardless of vitamin E supplementation. Also, irradiation reduced putrefying, and fishy odors during longer display times. TBARS increased with increased display time but was not affected by vitamin E supplementation. These results indicate that in aerobically packaged ground pork and vacuum packaged loin chops, radiolytic odors and color changes cannot be mediated with vitamin E supplementation. Display time and irradiation are significant in determining color and odor changes.

Community perception of school-based delivery of anthelmintics in Ghana and Tanzania.

This paper presents the results of an evaluation of community perception of two large-scale, government-run, school-based health programmes delivering anthelmintic drugs to primary school children, in Ghana (80 442 children in 577 schools) and Tanzania (110 000 children in 352 schools). Most teachers (96% in Ghana and 98% in Tanzania) were positive about their role in the programme, including administration of anthelmintic drugs, and parents and children fully accepted their taking on this role. The benefits of the programme were apparent to teachers, parents and children in terms of improved health and well-being of the children. Over 90% of parents in both Ghana and Tanzania indicated a willingness to pay for the continuation of drug treatment. The evaluation also highlighted areas that are critical to programme effectiveness, such as communication between schools and parents, the issue of collaboration between the health and education sectors, parents' perception of the importance of helminth infection as a serious and chronic health problem (compared with more acute and life threatening illnesses such as malaria), and who should pay for treatment of side-effects.

Validity of women's self-reported obstetric complications in rural Ghana.

This retrospective study assessed the utility of women's self-reports to identify obstetric complications in rural Ghana. All consenting obstetric and postpartum inpatients, presenting from the seventh month of gestation to 42 days postpartum, were interviewed at the Holy Family Hospital, Techiman and were asked about their signs and symptoms. A combination of clinical examination and laboratory testing of urine and blood samples was used for determining case status. Self-reported obstetric complications of 340 women were compared with the corresponding diagnostic status for their sensitivity, specificity, predictive value, and test-efficiency. Using algorithms that could not be practically applied at the community level, self-reported symptoms correctly identified the majority (75%) of complicated and uncomplicated pregnancies, but missed one-quarter of cases requiring emergency obstetric care. The positive predictive value of 50% indicates that women's self-reported symptoms should not be used in estimating the incidence of these conditions or in identifying women requiring referral in this population.

Continuous quality improvement in public health in Ghana: CQI as a model for primary health care management and delivery.

This article describes the context and examines factors influencing the quality of primary health care delivery and management in Ghana, West Africa. It describes the potential of continuous quality improvement as a management philosophy and tool to improve the quality of primary health care delivery and management in Ghana. It compares the Ghanaian context in which the use of continuous quality improvement in health care is new and untried with the developed country context (mainly the U.S.), where continuous quality improvement has a relatively long history of use in health care and is a requirement for the accreditation of health institutions. Finally, the article discusses the steps that have to be taken to translate continuous quality improvement from a theoretical management concept to improve quality of care to an actual managerial intervention in Ghana. In conclusion, continuous quality improvement is shown to be a potential viable approach to improving quality of care in the Ghanaian context and merits further investigation.

Impact of prepackaging antimalarial drugs on cost to patients and compliance with treatment.

OBJECTIVE: To examine the extent to which district health teams could reduce the burden of malaria, a continuing major cause of mortality and morbidity, in a situation where severe resource constraints existed and integrated care was provided. METHODS: Antimalarial drugs were prepackaged into unit doses in an attempt to improve compliance with full courses of chemotherapy. FINDINGS: Compliance improved by approximately 20% in both adults and children. There were 50% reductions in cost to patients, waiting time at dispensaries and drug wastage at facilities. The intervention, which tended to improve both case and drug management at facilities, was well accepted by health staff and did not involve them in additional working time. CONCLUSION: The prepackaging of antimalarials at the district level offers the prospect of improved compliance and a reduction in the spread of resistance.

Anaemia in schoolchildren in eight countries in Africa and Asia.

OBJECTIVE: To report on the haemoglobin concentrations and prevalence of anaemia in schoolchildren in eight countries in Africa and Asia. DESIGN: Blood samples were collected during surveys of the health of schoolchildren as a part of programmes to develop school-based health services. SETTING: Rural schools in Ghana, Indonesia, Kenya, Malawi, Mali, Mozambique, Tanzania and Vietnam. SUBJECTS: Nearly 14 000 children enrolled in basic education in three age ranges (7-11 years, 12-14 years and > or =15 years) which reflect the new UNICEF/WHO thresholds to define anaemia. RESULTS: Anaemia was found to be a severe public health problem (defined as >40% anaemic) in five African countries for children aged 7-11 years and in four of the same countries for children aged 12-14 years. Anaemia was not a public health problem in the children studied in the two Asian countries. More boys than girls were anaemic, and children who enrolled late in school were more likely to be anaemic than children who enrolled closer to the correct age. The implications of the four new thresholds defining anaemia for school-age children are examined. CONCLUSIONS: Anaemia is a significant problem in schoolchildren in sub-Saharan Africa. School-based health services which provide treatments for simple conditions that cause blood loss, such as worms, followed by multiple micronutrient supplements including iron, have the potential to provide relief from a large burden of anaemia.

Pharmacists' role in managing sexually transmitted infections: policy issues and options for Ghana.

The debate about extending the role of pharmacists in health care is growing in recognition of the ongoing difficulties experienced by many public sector services. The perceived accessibility and confidentiality of pharmacists makes them particularly attractive to patients for the management of health problems such as sexually transmitted infections (STI) that may lead to stigmatization. Despite growing interest in the subject, there are few documented analyses of the role of pharmacists in low-income countries. In Ghana, pharmacists are acknowledged by the government to be the preferred option for people seeking treatment for STI. A study was conducted to investigate the current role played by pharmacists in Greater Accra Region in the management of STI. On the basis of these findings, training schemes were developed, implemented and evaluated. This paper presents the findings of this operation-research and considers their implications for deciding to what extent and in what way pharmacists should be involved in managing STI in Ghana and other similar country settings. These findings suggest that pharmacists have a crucial role in effective management of STI, particularly in the management of urethral discharge. They may need to limit their management of genital ulcer to referring customers to laboratories and medical practitioners. They also represent a currently under-utilized opportunity for preventive activities. Regulation and quality assurance issues need to be addressed by both pharmacy and medical professions.

The effect of recall on estimation of incidence rates for injury in Ghana.

BACKGROUND: Injury is a major public health problem in many developing countries. Due to limitations of vital registry and health service data, surveys are an important tool to obtain information about injury in these countries. The value of such surveys can be limited by incomplete recall. The most appropriate recall period to use in surveys on injury in developing countries has not been well addressed. METHODS: A household survey of injury in Ghana was conducted. Estimated annual non-fatal injury incidence rates were calculated for 12 recall periods (1-12 months prior to the interview, with each successively longer period including the preceding shorter periods). RESULTS: There was a notable decline in the estimated rate from 27.6 per 100 per year for a one-month recall period to 7.6 per 100 per year for a 12-month recall period (72% decline). The extent of this decline was not influenced by age, gender, rural versus urban location, nor by type of respondent (in-person versus proxy). Rate of decline was influenced by severity of injury. Injuries resulting in <7 days of disability showed an 86% decline in estimated rates from a one-month to a 12-month recall period, whereas injuries resulting in > or =30 days of disability showed minimal decline. CONCLUSIONS: In this setting, longer recall periods significantly underestimate the injury rate compared to shorter recall periods. Shorter recall periods (1-3 months) should be used when calculating the overall non-fatal injury incidence rate. However, longer recall periods (12 months) may be safely used to obtain information on the more severe, but less frequent, injuries.