Sero-epidemiology of toxoplasmosis amongst pregnant women in the greater accra region of ghana.

OBJECTIVES: To investigate Toxoplasma infection among pregnant women in relation to exposure to infection risk, age and pregnancy-related risk factors. DESIGN AND METHODS: This cross-sectional study involved 294 pregnant women attending ante-natal clinic in Accra who consented to participate. Personal and Toxoplasma infection risk related data were obtained by questionnaire interviews. Venous blood was safely drawn from each participant and spun to obtain sera. Each of the 159 randomly selected serum samples was tested for specific anti-Toxoplasma (anti-T. gondii) antibodies IgG, IgA and IgM using a commercial ELISA kit (Calbiotech Inc., CA). ELISA results were correlated with exposure to possible infection risk factors as well as age and pregnancy-related risk factors. RESULTS: The 159 women aged 15-40 years in their first, second and third trimesters, numbered 29, 70 and 60, respectively. An overall anti-T. gondii antibodies IgG, IgA and IgM seroprevalence of 92.5% (147/159) was recorded, with 4.1% (6/147) of them having anti-IgG only. The remaining 88.7% (141/159) had anti-Toxoplasma antibodies IgG, IgA and IgM in various combinations and consisted of 17.7% (25/141) in their first, 44.0% (62/141) in their second, and 38.3% (54/141) in their third, trimesters. Twelve women (7.6%) were seronegative for all 3 antibodies CONCLUSIONS: Seroprevalence was high among the women and exposure to contact with cats' faeces was found to be the major T. gondii infection risk factor. Age and pregnancy-related risk factors did not have association with T. gondii infection within the limitations of this study.

Experimental control of Biomphalaria pfeifferi, the intermediate host of Schistosoma mansoni, by the ampullariid snail Lanistes varicus.

The biological control of the snail hosts of the trematodes that cause human schistosomiasis appears to be a promising method for achieving sustainable reductions in the transmission of the parasites. The possibility of using the Ghanaian strain of an ampullariid snail, Lanistes varicus, for the biological control of the main snail host of Schistosoma mansoni , Biomphalaria pfeifferi, has now been investigated in laboratory-based experiments. Adult and 2-week-old L. varicus were found to feed voraciously on the egg masses and juveniles of B. pfeifferi (from the Tono irrigation canals in northern Ghana). When single L. varicus were exposed to 20-200 egg masses, they consumed all of the masses over 24 h (if adult) or about 50% of them over 4 days (if 2-week-old juveniles). The effect of the secretions of the ampullariid on the reproduction, growth and mortality of B. pfeifferi was also investigated, by maintaining the two snail species in the same aquarium but separated by nylon netting. The presence of L. varicus in the same aquarium reduced the number of egg masses produced by each B. pfeifferi, although, curiously, the presence of a single L. varicus in the aquarium appeared to have more of an impact, on the egg-mass deposition by 20 B. pfeifferi, than the presence of five or more of the ampullariids. It appears that, under laboratory conditions at least, the Ghanaian stain of L. varicus has the potential to limit populations of B. pfeifferi.

Multiple Plasmodium falciparum infections in Tanzanian infants.

Paired blood samples from 99 Tanzanian infants were analysed to examine the infection dynamics of Plasmodium falciparum during the first year of life. Infecting parasites were genotyped by polymerase chain reaction amplification of the polymorphic gene for the merozoite surface protein 2 and subsequent analysis according to the resulting restriction fragment length polymorphism pattern. The same samples served as controls in a parallel case-control study for which an additional blood sample was taken from each child during a fever episode. The relationship of the number of concurrent infections (multiplicity) with age and morbidity was analysed and results were compared to those of a similar study on older children between 2 and 7 years of age, carried out in the same village at the same time. The mean of 2 infecting genotypes per positive blood sample from community surveys was low compared to that in older children, and there was no significant age-dependency of multiplicity within the first year of life. Multiplicity of infection in fever cases was also independent of age. In infants, multiplicity was positively associated with parasite density and risk of clinical malaria, in contrast to the situation in older children (> 2 years). The findings help in the understanding of infection dynamics, premunition, and development of semi-immunity in malaria.

Effect of insecticide-treated bed nets on haemoglobin values, prevalence and multiplicity of infection with Plasmodium falciparum in a randomized controlled trial in Tanzania.

A randomized controlled trial of insecticide-treated bed nets (ITNs) was conducted in an area of high malaria transmission in Tanzania in order to assess the effects of ITNs on infection and anaemia. One hundred and twenty-two children, aged 5 to 24 months, were randomly allocated to 2 groups, one of which received ITNs. Outcome measures were assessed in 6 consecutive months with monthly cross-sectional surveys. These measures were haemoglobin values, Plasmodium falciparum prevalence and density, and multiplicity of infection determined by polymerase chain reaction-restriction fragment length polymorphism analysis (PCR-RFLP) of the msp2 locus. There was a significant increase in mean heamoglobin values and a significant decrease of 16.4% in microscopically determined P. falciparum prevalence in children in the ITN group six months after the start of the trial. Both effects were more pronounced in younger children. However, no significant difference was observed in parasite density or multiplicity of infection among infected children. Comparison with PCR results indicated that microscopically subpatent parasitaemia was more frequently found in children in the ITN group. This, together with the observed similar multiplicity in the 2 groups, suggests that infections are maintained despite ITN use, owing to the chronicity of infections. This study shows that ITNs reduce the risk of anaemia in highly exposed young children. The virtually unchanged multiplicity of infection indicates that the potentially protective concomitant immunity is not compromised.

Pyrimethamine-resistant Plasmodium falciparum parasites among Tanzanian children: a facility-based study using the polymerase chain reaction.

A mutation-specific polymerase chain reaction method was used to estimate the proportion of pyrimethamine-resistant parasites in 101 children reporting with malaria at the hospital in Ifakara, a town in southern Tanzania. The method is based on the observation that a point mutation (Asn-108) in the dihydroifolate reductase gene confers resistance to pyrimethamine. Twenty-eight percent of the examined 101 children had pyrimethamine-resistant parasites, 65% had pyrimethamine-sensitive parasites with the wild-type Ser-108 codon, and 9% had both alleles, suggesting a mixed infection. None of the 21 children with clinical malaria had pyrimethamine-resistant parasites. Currently, sulfadoxine-pyrimethamine is considered a potential first-line drug for malaria treatment in most African countries. We suggest that although sulfadoxine-pyrimethamine could still be effective against chloroquine-resistant malaria in this area, its judicious use is important so as to minimize the spread of resistance.

PCR amplification of DNA from malaria parasites on fixed and stained thick and thin blood films.

Under some circumstances, polymerase chain reaction (PCR) amplification of deoxyribonucleic acid (DNA) from Plasmodium may become necessary from infections for which only blood slides are available. Established methods used for DNA preparation do not work in that case. We have developed a reliable and controlled method for DNA preparation from malaria parasites on fixed and stained blood films. 162 slides from 2 different locations, some stored for at least one year, have been analysed by PCR amplification of the polymorphic loci for MSA1 and MSA2. In 92% of microscopically positive slides, a PCR product could be detected using material derived from thick blood films. When thin blood films with scanty parasitaemia were used, a PCR product could be obtained with only 71% of samples. In all unsuccessful cases, DNA preparation was the limiting factor, which was controlled for by amplification of a control human template.

Dynamics of pyrimethamine-resistant P. falciparum parasites in sulphadoxine-pyrimethamine treated Tanzanian children.

A mutation -specific PCR assay was used to determine the dynamics of pyrimethamine-resistant P. falciparum parasites in sulphadoxine-pyrimethamine treated children, 0-5 years, from a village in Tanzania. The assay is based on the observation that a point mutation in the dihydrofolate reductase (DHFR) gene confers resistance to pyrimethamine. The PCR assay was used on blood samples collected from treated children on days 0,2,7,14,21and 28. Preliminary results revealed that pyrimethamine-sensitive parasites seemed to be completely cleared after 7 or 14 days of treatment but re-surfaced after 21 days. It was observed that in those children without mixed infection on day 0, pyrimethamine-resistant parasites appeared after 7 days. The implifications of these findings are discussed.

Dynamics of Pyrimethamine-Resistant P. Falciparum parasites in Sulphadoxine-Pyrimethamine Treated in Children

A mutation -specific PCR assay was used to determine the dynamics of pyrimethamine-resistant P. falciparum parasites in sulphadoxine-pyrimethamine treated children; 0-5 years; from a village in Tanzania. The assay is based on the observation that a point mutation in the dihydrofolate reductase (DHFR) gene confers resistance to pyrimethamine. The PCR assay was used on blood samples collected from treated children on days 0;2;7;14;21and 28. Preliminary results revealed that pyrimethamine-sensitive parasites seemed to be completely cleared after 7 or 14 days of treatment but re-surfaced after 21 days. It was observed that in those children without mixed infection on day 0; pyrimethamine-resistant parasites appeared after 7 days. The implifications of these findings are discussed

Polymerase chain reaction used to describe larval habitat use by Anopheles gambiae complex (Diptera: Culicidae) in the environs of Ifakara, Tanzania.

Larvae of the Anopheles gambiae complex were collected in and around the town of Ifakara, southern Tanzania during the wet season of 1994 and identified to species by polymerase chain reaction. All but 1 surface pool contained mixed populations of An. gambiae and An. arabiensis larvae. The 2 species varied among locations rather than types of water. An. arabiensis predominated in pools close to cattle. The numbers of identified early instars of both species were similar, but more An. gambiae 4th instars were identified, perhaps indicating that An. gambiae were able to survive heavy rainfall better than A. arabiensis.

Does chloroquine treatment influence the distribution of pyrimethamine-resistant P. falciparum parasites in the field?

In a limited case controlled but molecular study in symptomatic Tanzanian children, we found that those who had not previously received Chroloquine treatment had a higher proportion of parasitaemia. There was also a higher proportion of pyrimetamine-resistant P. falciparum parasites among those who had not received any treatment. This finding is paradoxical and the reasons for this observation are suggested.

Activity of U-snRNA genes with modified placement of promoter elements in transfected protoplasts and stably transformed tobacco.

In higher plants the promoter elements of pol II- and pol III-transcribed U-snRNA genes are identical, comprising a -30 TATA box and an upstream sequence element, USE. The USE and TATA are centred approximately four and three helical DNA turns apart in pol II and pol III genes, respectively, and it is this difference in the element spacing that determines the RNA polymerase specificity of the gene. In this study we have analyzed the effect of spacing mutations on activity of Arabidopsis U2 and U6 genes in transfected protoplasts of Nicotiana plumbaginifolia and in stably transformed tobacco. In the pol III-transcribed U6 gene the insertions and deletions of either odd or even numbers of half helical turns completely inactivate transcription in transfected protoplasts, consistent with the high conservation of the element spacing found in all plant U-snRNA genes. Surprisingly, while insertions of 50 base pairs (bp) or more into the spacer of the pol II-specific U2 gene inactivate transcription, a deletion of 5 bp or insertions of as much as 20 bp decrease transcription by only 40 to 70%. This relaxed requirement for the conserved element spacing is only seen in transfected protoplasts since the same mutant U2 genes are not transcribed in stably transformed tobacco when transcription takes place from the chromosome. The results provide some clues about possible factor interactions at the promoters of plant U-snRNA genes and also offer an example of major differences in transcription between transiently and stably transformed cells.

Structure and expression of the U5 snRNA gene of Arabidopsis thaliana. Conserved upstream sequence elements in plant U-RNA genes.

We have previously characterized the U2 small nuclear (sn) RNA gene family of Arabidopsis thaliana. To find out the structural features of upstream and downstream non-coding regions that are shared by different U-RNA genes in higher plants we have isolated the gene encoding a 125 nt-long U5 snRNA of Arabidopsis. Activity of the cloned gene was demonstrated in stably transformed tobacco calli and by transient expression in transfected protoplasts of Nicotiana plumbaginifolia. Southern analysis indicated that the Arabidopsis genome contains 8-9 copies of the U5 gene. Alignment of upstream non-coding regions revealed two elements conserved between all plant U-RNA genes characterized so far: the sequence RTCCCACATCG (-70/-80 region, 100% conservation) and the TATA homology around position -30. The coding regions in all genes are followed by the sequence CAN4-9AGTN (A/T)AA which may correspond to a termination and/or processing signal.