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Pavonia odorata Willd. belonging to the family Malvaceae is well known for curing a variety of diseases. This study deals with the Pharmacognostical and phytochemical evaluation of whole plant of Pavonia odorata Willd. The plant is widely used as a cooling, diaphoretic, diuretic, and demulcent. It is also prescribed as an astringent and tonic for fever, inflammation and hemorrhage. Pharmacognostical studies include macroscopic, microscopic and physicochemical evaluation of the whole plant. The transverse section of root, stem and leaves were performed. Physicochemical constants showed the values of total ash (4.45%), acid insoluble ash (0.754%), water soluble ash (6%), sulphated ash (6. 5%), nitrated ash (1.65%), hot extraction(10.22%), water soluble extractive (9.2%), ethanol soluble extractive (4.8%), ether soluble extractive (1.2%), loss on drying (25%), swelling index(1) and mucilage content(0.25%) w/w. The preliminary phytochemical screening of the alcoholic extract showed presents of alkaloids, phenolic compounds, steroids, mucilage and flavonoids. The study helps in the development of standard parameters of the plant which help the identification and authentication of the plant. Keywords: Pavonia odorata Willd., Pharmacognostical, phytochemical evaluation, Medicinal plant drugs, ethnomedicine.

Introduction: Around 5,00,000 women are affected by cervical cancer and nearly half of them end up losing the battle of life with this deadly disease. So, there is an urgent need for the synthesis and development of new, small, synthetic molecules to tackle this challenge. Schiff’s bases are derivatives of azomethine group (-CH=N-) and are highly reactive. Here a series of novel Schiff’s bases were synthesized by single step process of condensing substituted 2-amino benzothiazole with different benzaldehydes. Objectives: To design, synthesize novel Schiff’s bases of 2-amino benzothiazole and evaluate its anti-oxidant as well as anti-cancer activity. Methods: A total of 18 compounds were synthesized by single step process of condensing substituted 2-amino benzothiazole with different substituted benzaldehydes. These were characterized by FTIR, 1H NMR, and Mass spectroscopy. The synthesized compounds were tested in-vitro for both antioxidant and antiproliferative activity. In-silico docking studies were performed on the crystal structure of the complex of caspase-3 with a nicotinic acid aldehyde inhibitor with PDB IDs 1RE1, 1RHM and 3DEH to study the interaction of the compounds with the receptor. Results: Majority of the derivatives displayed moderate to significant antiproliferative activity on HeLa cell line. Interestingly, the compound SP16 showed excellent activity with an IC50 value of 2.517μg/ml in comparison to the reference compound Cisplatin (17.2μg/ml). Compound SP7 and SP 15 showed favourable in silico interactions.Conclusion: A series of 18 novel Schiff’s bases of 2-amino benzothiazoles compounds were designed, synthesized and evaluated for their biological activities. The compound SP16 showed excellent activity with an IC50 value of 2.517μg/ml in comparison to the reference compound Cisplatin and Compound SP7 and SP 15 showed favourable in silico interactions.

The epidemiology of giardiasis in rural villages in Peninsular Malaysia was examined in the context of the One Health triad that encompasses humans, animals and environment (i.e. river water). A cross-sectional study was carried out among five rural communities in Malaysia to determine the prevalence of Giardia duodenalis in humans, animals and river water. Fecal samples collected from humans and animals were examined by light microscopy. Water was sampled from the rivers adjacent to the target communities and investigated for the occurrence of Giardia cysts. The isolated cysts were further genotyped targeting the glutamate dehydrogenase and triosephosphate isomerase genes. The overall prevalence of G. duodenalis was 6.7% (18/269) and 4.7% (8/169) among humans and animals, respectively. Giardia cysts (mean concentration range: 0.10-5.97 cysts/L) were also found in adjacent rivers at four out of the five villages examined. At Kemensah and Kuala Pangsun, Giardia cysts were isolated from humans [rate: 3.7% each (of 54 each)], animals [rates: 6.3% (of 62) and 11.3% (of 16), respectively] and river water [average concentration of 9 samples each: 0.83±0.81 and 5.97±7.00, respectively]. For both villages at Pos Piah and Paya Lebar, 12.2% (of 98) and 6.1% (of 33) of collected human samples were infected, respectively whilst none of the collected animals samples in these villages were found to be positive. The river water samples of these two villages were also contaminated (average concentration: 0.20±0.35 (of 9) and 0.10±0.19 (of 3), respectively). In conclusion, Giardia cysts were simultaneously observed in the human-animal-environment (i.e., river water) interfaces in at least two of five studied communities highlighting a vital need to improve understanding on the interplay of transmission dynamics, the role of infected humans and animals in contaminating the water sources and the role of water as a vehicle of disease transmission in these communities. Indeed, this study illustrates the One Health approach which is to recognize that the optimal health of humans are interconnected with the well-being of animals and their environment.

Yemen is a developing country overwhelmed with a triad of poverty, diseases and social conflicts. Moreover, the majority of its population live in rural communities and suffer from intestinal parasitic infections (IPIs). Therefore, the present school-based, cross-sectional survey aimed to detect the prevalence of such infections and associated risk factors among schoolchildren in the rural communities of Bani Alharith, Hamdan and Bani Hushaysh districts of Sana'a, north of Yemen. Socio-demographic data and certain behavioral risk factors as well as stool samples were collected from 1218 schoolchildren from ten randomly schools in the study area. Fresh stool samples were examined for parasites by direct saline and iodine preparations and after concentration with formol-ether technique. The overall prevalence of IPIs was 54.8%, with a higher frequency of protozoal than helminthic infections (37.6 vs. 17.2%, respectively). Parasite species recovered were Entameba histolytica (21.5%), Giardia lamblia (16.1%), Ascaris lumbricoides (8.3%), Hymenolepis nana (5.3%), Schistosoma mansoni (2.6%), Trichuris trichiura (0.5%) and Enterobius vermicularis (0.4%). Univariate analysis showed that the male gender and illiteracy of fathers and/or mothers were the socio-demographic factors significantly associated with higher infection rates. The illiteracy of mothers was also confirmed as an independent risk factor by multivariable analysis. On the other hand, not washing hands before eating, not washing fruits and vegetables before consumption, eating uncovered food and not clipping fingernails were the risk behaviors significantly associated with higher infection rates, with the last three ones being confirmed as independent risk factors. Therefore, control measures should include regular treatment of protozoal infections and deworming of schoolchildren, promotion of hygiene in rural schools through health education programs, regular inspection of schoolchildren for personal hygiene practices and the provision of a healthy school infrastructure.

Since 2005, artesunate (AS) plus sulfadoxine/pyrimethamine (SP) combination has been adopted as the first-line treatment for uncomplicated malaria in Yemen in response to the high level of Plasmodium falciparum resistance to chloroquine (CQ). Therefore, the aim of the present study was to determine the frequency distribution of molecular markers associated with resistance to CQ and AS plus SP combination among P. falciparum isolates from a malaria-endemic area in Taiz governorate, Yemen. Fifty P. falciparum isolates were collected during a cross-sectional study in Mawza district, Taiz, in the period from October 2013 to April 2014. The isolates were investigated for drug resistance-associated molecular markers in five genes, including P. falciparum CQ resistance transporter (pfcrt) 76T and P. falciparum multidrug resistance 1 (pfmdr1) 86Y as markers of resistance to CQ, mutations in the Kelch 13 (K13) propeller domain for resistance to AS, and P. falciparum dihydrofolate reductase (pfdhfr) and P. falciparum dihydropteroate synthase (pfdhps) genes for resistance to SP. Nested polymerase chain reaction was used to amplify target genes in DNA extracts of the isolates followed by restriction fragment length polymorphism for detecting 76T and 86Y mutations in pfcrt and pfmdr1, respectively, and by DNA sequencing for detecting mutations in K13, pfdhfr and pfdhps. All the investigated isolates from Mawza district were harboring the pfcrt 76T mutant and the pfmdr1 N86 wild-type alleles. The pfdhfr 51I/108N double mutant allele was found in 2.2% (1/45) of the isolates; however, no mutations were detected at codons 436, 437, 540, 581 and 613 of pfdhps. All P. falciparum isolates that were successfully sequenced (n=47) showed the K13 Y493, R539, I543 and C580 wild-type alleles. In conclusion, the pfcrt 76T mutant allele is fixed in the study area about six years after the official withdrawal of CQ, possibly indicating its over-the-counter availability and continued use as a self-medication in the study area. However, the almost predominant wild-type alleles of the genes associated with resistance to AS and SP among P. falciparum isolates in the present study indicates the sustained efficacy of the currently adopted first-line treatment of AS plus SP in the study area.

Onchocerciasis is a neglected parasitic disease affecting the poorest underserved people in Yemen. A national control programme with goals to eliminate onchocerciasis has yet to be launched due to the current upheaval and social unrest in the country. The disease, locally termed as sowda, is unique in its clinicopathologic pattern, being of the localized, non-blinding, hyperreactive onchocercal skin disease. Although early reports identified endemic foci along seasonal watercourses, there is a need to redefine its epidemiologic patterns as well as health and socioeconomic impacts. Laboratory diagnosis of sowda among Yemeni patients is difficult due to the low load of microfilariae in skin snips and the presence of asymptomatic itching-free microfilaria carriers. Adoption of ivermectin use at three-month intervals as a control strategy has not been evaluated because the drug is mostly used in clinics and distributed to only a few affected communities. This paper addresses key aspects of onchocerciasis in Yemen and highlights the need for screening at-risk populations using highly sensitive techniques and mapping the distributions of the parasite in human and vector populations of blackflies. The new research should be integrated with the launch of a national onchocerciasis control programme to achieve onchocerciasis elimination.

Background Onchocerciasis in Yemen is one of the most neglected diseases, where baseline estimates of onchocerciasis and monitoring of the impact of ivermectin regularly administered to the affected individuals on its transmission are lacking. Therefore, this study aimed to determine the anti-Ov16 IgG4 seroprevalence among local communities of Hodeidah and Al-Mahwit governorates of Tihama region. The factors possibly associated with previous exposure to infection were also studied. Methodology/Principal findings This cross-sectional study was conducted in two ivermectin-targeted districts endemic for onchocerciasis in Hodeidah and Al-Mahwit and two untargeted districts with unknown previous endemicity in Hodeidah between February and July 2017. For 508 residents sampled by a multi-stage random approach, data were collected and blood specimens were screened for anti-Ov16 IgG4 using the SD BIOLINE Onchocerciasis IgG4 rapid tests. The study revealed an overall anti-Ov16 IgG4 rate of 18.5% (94/508) in all surveyed districts, with 10.2% (12/118) of children aged ≤10 years being seropositive. Moreover, rates of 8.0% (4/50) and 6.1% (4/66) were found in districts not officially listed as endemic for the disease. Multivariable analysis confirmed the age of more than ten years and residing within a large family as the independent predictors of exposure to infection. Conclusions/Significance Onchocerciasis transmission is still ongoing as supported by the higher anti-Ov16 IgG4 seroprevalence rate among children aged ≤10 years compared to that (, Author summary Onchocerciasis is endemic in certain foci in the western governorates of Yemen. Monitoring the impact of the regular ivermectin administration to affected individuals on the transmission status and providing baseline onchocerciasis estimates in endemic areas are crucial for planning effective elimination strategies. We found that the disease transmission is still ongoing in Hodeidah and Al-Mahwit governorates of Tihama region as indicated by the anti-Ov16 IgG4 seropositivity among children aged ≤10 years. In Bani Sa'ad, where affected individuals had been regularly targeted with ivermectin over the last 15 years, we found that the anti-Ov16 IgG4 seroprevalence rate was significantly lower among children aged ≤10 years (9.1%; 5/55) compared to those >10 years (24.5%; 37/151), reflecting a possible decline in disease transmission. We also revealed onchocerciasis transmission in districts with unknown previous endemicity for the first time, with rates of 8.0% and 6.1% being found in Al Marawi'ah and As Sukhnah districts of Hodeidah. Large-scale surveys are recommended for mapping of O. volvulus in human and blackfly populations in endemic foci and neighboring untargeted areas of uncertain endemicity as a forward step towards the elimination of the disease from the country.

Background Toxoplasma gondii is a zoonotic coccidian parasite causing morbidity and mortality. In Yemen, T. gondii infection has been reported among pregnant women seeking healthcare in the main cities. However, no data are available on the prevalence of T. gondii infection and its associated risk factors among pregnant women in the rural communities of the country. Thus, the present study aimed to determine the seroprevalence of T. gondii and identify its risk factors among pregnant women in the rural communities of Taiz governorate, Yemen. Methods A total of 359 pregnant women living in the rural communities of Taiz governorate were enrolled in this study by house-to-house visits. Data were collected using a pre-designed questionnaire, and blood samples were collected and tested for the detection of anti- T. gondii IgM and IgG antibodies by enzyme-linked immunosorbent assay. Results The prevalence of T. gondii infection among pregnant women in this study was 46.2% (166/359). Bivariate analysis identified the age of ≥ 30 years (odds ratio [OR] = 1.7; 95% confidence interval [CI] = 1.09–2.65, P = 0.019) and unimproved water sources (OR = 2.2; 95% CI = 1.10–4.55, P = 0.023) as factors associated with T. gondii infection among pregnant women. The multivariable analysis, however, identified unimproved water sources as an independent risk factor (adjusted OR = 2.4; 95% CI = 1.16–5.0, P = 0.018) associated with T. gondii infection among pregnant women. Conclusions Pregnant women in the rural communities of Taiz, Yemen are at high risk of contracting T. gondii infection. Unimproved water sources (wells, water streams and water tanks) are significantly associated with T. gondii infection and should be considered in prevention and control strategies, especially among pregnant women. Electronic supplementary material The online version of this article (doi:10.1186/s40249-017-0243-0) contains supplementary material, which is available to authorized users.

Background In the Arabian Peninsula malaria control is progressing steadily, backed by adequate logistic and political support. As a result, transmission has been interrupted throughout the region, with exception of limited sites in Yemen and Saudi Arabia. Here we examined Plasmodium falciparum parasites in these sites to assess if the above success has limited diversity and gene flow. Methods We examined 108 P. falciparum isolates in three sites in Yemen (Taiz, Dhamar and Hodeidah) and 91 isolates from Saudi Arabia (Jazan). Nine microsatellites were analyzed for allelic diversity, multi-locus haplotype and inter-population differentiation. Results Diversity at each locus (unbiased heterozygosity [H]) was relatively lower in Yemen; (Hodeidah, H = 0.615, Taiz, H = 0.66, Dhamar, H = 0.481), compared to Saudi Arabia (Jazan, H = 0.76). Microsatellites were distributed widely and private alleles, detected in a single population, were rare. Pairwise comparisons revealed that parasites population in Dhamar was relatively distanced (FST = 0.19). However, Taiz (Yemen) (FST = 0.065) and Hodeidah (FST = 0.107) populations were closer to that in Jazan (Saudi Arabia). Nonetheless, parasites in the four sites can be considered as one population. Conclusion Although malaria risk in Saudi Arabia has been cut considerably, the extent of diversity and parasite genetic structure are indicative of a large population size. Elimination strategy should target demographic factors that favor parasite dispersal and flow of imported malaria.

Population displacement and other forced movement patterns following natural disasters, armed conflicts or due to socioeconomic reasons contribute to the global emergence of Aedes-borne viral disease epidemics. In particular, dengue epidemiology is critically affected by situations of displacement and forced movement patterns, particularly within and across borders. In this respect, waves of human movements have been a major driver for the changing epidemiology and outbreaks of the disease on local, regional and global scales. Both emerging dengue autochthonous transmission and outbreaks in countries known to be non-endemic and co-circulation and hyperendemicity with multiple dengue virus serotypes have led to the emergence of severe disease forms such as dengue hemorrhagic fever and dengue shock syndrome. This paper reviews the emergence of dengue outbreaks driven by population displacement and forced movements following natural disasters and conflicts within the context of regional and sub-regional groupings.

This study aims to assess the bactericidal activity of Propolis extract against methicillin resistance Staphylococcus aureus (MRSA) in vivo. It included (11 male white New Zealand rabbits) evenly divided into two groups (6 animals in each one). Group I was fed daily with one milliliter of 20% extract Propolis for twenty days, while group II was fed without Propolis to serve as a control group. Ten days after the last dose, all animals were injected in intraperitoneal with single dose viable MRSA antigen at 0.5 x108 cfu/ml. After two days, blood samples that collected from all animals by heart puncture were cultured on blood agar media to assess bactericidal power activity of extract Propolis. The results pointed to effect of this extract on survive viable MRSA bacteria as compared with survive of same bacteria in control group animals. It was concluded Propolis had bactericidal power against pathogenic bacteria that appear resistant to antibiotics like MRSA bacteria.

Giardia duodenalis is an important intestinal protozoan in Yemen with infection rates ranging from 18% to 27%. To date, there has been no genotyping study to provide a better understanding of the transmission dynamic. This study was conducted to genotype and subtype G. duodenalis in Yemen. Stool samples were collected from 503 Yemeni outpatients between 1 and 80 years old, including 219 males and 284 females. Giardia cysts were detected via microscopy after the formal-ether concentration. Genotyping of Giardia was carried out using PCR and sequence analysis of the 16s rRNA and b-giardin genes. Of the 89 microscopy-positive Giardia samples, 65 were successfully sequenced, of which 66% (43 of 65) were identified as G. duodenalis assemblage A and 34% (22 of 65) as assemblage B. Further subtyping analysis based on b-giardin gene identified the presence of subtypes A2 and A3, which belong to the anthroponotic sub-assemblage AII. Data of the study suggest that anthroponotic transmission played a potential role in the transmission of giardiasis in the community. However, further genotyping and subtyping studies of specimens from humans and animals living in the same households are needed for a more definitive understanding of giardiasis transmission in Yemen.

SUMMARYCryptosporidiumis a protozoan parasite of humans and animals and has a worldwide distribution. The parasite has a unique epidemiology in Middle Eastern countries where the IId subtype family ofCryptosporidium parvumdominates. However, there has been no information onCryptosporidiumspecies in Yemen. Thus, this study was conducted in Yemen to examine the distribution ofCryptosporidiumspecies and subtype families. Fecal samples were collected from 335 patients who attended hospitals in Sana'a city.Cryptosporidiumspecies were determined by PCR and sequence analysis of the 18 s rRNA gene.Cryptosporidium parvumandC. hominissubtypes were identified based on sequence analysis of the 60 kDa glycoprotein (gp60) gene. Out of 335 samples, 33 (9·9%) were positive forCryptosporidium. Of them, 97% were identified asC. parvumwhilst 1 case (3%) was caused byC. hominis. All 7C. parvumisolates subtyped belonged to the IIaA15G2R1 subtype. The common occurrence of the zoonotic IIa subtype family ofC. parvumhighlights the potential occurrence of zoonotic transmission of cryptosporidiosis in Yemen. However, this postulation needs confirmation with future molecular epidemiological studies of cryptosporidiosis in both humans and animals in Yemen.

Malaria in Yemen is mainly caused by Plasmodium falciparum and 25 % of the population is at high risk. Sulfadoxine–pyrimethamine (SP) had been used as monotherapy against P. falciparum. Emergence of chloroquine resistance led to the shift in anti-malarial treatment policy in Yemen to artemisinin-based combination therapy, that is artesunate (AS) plus SP as first-line therapy for uncomplicated malaria and artemether–lumefantrine as second-line treatment. This study aimed to screen mutations in the dihydrofolate reductase (dhfr) and dihydropteroate synthetase (dhps) genes associated with SP resistance among P. falciparum population in Hadhramout governorate, Yemen. Genomic DNA was extracted from dried blood spots of 137 P. falciparum isolates collected from a community-based study. DNA was amplified using nested polymerase chain reaction (PCR) and subsequently sequenced for Pfdhfr and Pfdhps genes. Sequences were analysed for mutations in Pfdhfr gene codons 51, 59, 108, and 164 and in Pfdhps gene codons 436, 437, and 540. A total of 128 and 114 P. falciparum isolates were successfully sequenced for Pfdhfr and Pfdhps genes, respectively. Each Pfdhfr mutant allele (I 51 and N 108 ) in P. falciparum population had a frequency of 84 %. Pfdhfr R 59 mutant allele was detected in one isolate. Mutation at codon 437 (G 437 ) in the Pfdhps gene was detected in 44.7 % of falciparum malaria isolates. Frequencies of Pfdhfr double mutant genotype (I 51 C59 N 108 I164) and Pfdhfr/Pfdhps triple mutant genotype (I 51 C59 N 108I164-S436 G 437K540) were 82.8 and 39.3 %, respectively. One isolate harboured Pfdhfr triple mutant genotype (I 51 , R 59 , N 108 , I164) and Pfdhfr/Pfdhps quadruple mutant genotype (I 51 R 59 N 108I164-S436 G 437K540). High frequencies of Pfdhfr and Pfdhps mutant alleles and genotypes in P. falciparum population in Hadhramout, Yemen, highlight the risk of developing resistance for SP, the partner drug of AS, which subsequently will expose the parasite to AS monotherapy increasing then the potential of the emergence of AS resistance. Study findings necessitate the continuous monitoring of the efficacy of the national anti-malarial drugs policy in Yemen. In addition, monitoring SP efficacy using molecular markers that has shown to be a practical and informative method for monitoring the partner drug of AS.

SUMMARYCryptosporidiumspecies are protozoan parasites that infect humans and a wide variety of animals. This study was aimed at identifyingCryptosporidiumspecies and genotypes isolated from avian hosts. A total of 90 samples from 37 different species of birds were collected throughout a 3-month period from April 2008 to June 2008 in the National Zoo of Kuala Lumpur, Malaysia. Prior to molecular characterization, all samples were screened forCryptosporidiumusing a modified Ziehl-Neelsen staining technique. Subsequently samples were analysed with nested-PCR targeting the partial SSU rRNA gene. Amplicons were sequenced in both directions and used for phylogenetic analysis using Neighbour-Joining and Maximum Parsimony methods. Although 9 (10%) samples were positive forCryptosporidiumvia microscopy, 8 (8·9%) produced amplicons using nested PCR. Phylogenetic trees identified all the isolates asCryptosporidium parvum. AlthoughC. parvumhas not been reported to cause infection in birds, and the role of birds in this study was postulated mainly as mechanical transporters, these present findings highlight the significant public health risk posed by birds that harbour the zoonotic species ofCryptosporidium.

Ethyl acetate extracts of fermentation cultures of five local Streptomyces isolates SRY-3, SRY-25, 5b, 10, NS-38 were identified as inhibitors of plasmacytoma cell line. Concentration at 0.016 µg/ml of ethyl acetate extract of (SRY-3,SRY-25,5b,10,NS-38) inhibited (46%,40%,21%,34%,34%) of mouse plasmacytoma cells respectively, where as 0.5 µg/ml inhibited (58%, 55%, 35%, 33%, 36%) of plasmacytoma cells .Quantities analysis of crude extracts by using HPLC on the basis of their retention times showed that the values were (4.3,4.4,4.86,4.83&4.84) min. for SRY-3, SRY-25, 5b, 10,NS-38 respectively, while the retention time of standard antitumor compounds was (4.26) min. This suggests that unknown compound of SRY-3 extract contain bleomycin-like compound. Addition of standard bleomycin to crude extract of SRY-3 increased milliabsorbans unit (M.A.U.) from 0.49 to 1.4. The concentration of bleomycin-like antitumor in fermentation broth of SRY-3 isolate was 3.441 µg/ml.The five Streptomyces isolates have moderate activity against gram-positive and gram-negative bacteria, while standard BLM have no activity.

International audience; Standard anti‑malarial drug efficacy and drug resistance assessments neglect the gametocyte parameters in their pro‑ tocols. With the spread of drug resistance and the absence of clinically proven vaccines, the use of gametocytocidal drugs or drug combinations with transmission‑blocking activity is a high priority for malaria control and elimination. However, the limited repertoire of gametocytocidal drugs and induction of gametocytogenesis after treatment with certain anti‑malarial drugs necessitate both regular monitoring of gametocytocidal activities of anti‑malarial drugs in clinical use and the effectiveness of candidate gametocytocidal agents. Therefore, updating current protocols of anti‑ malarial drug efficacy is needed to reflect the effects of anti‑malarial drugs or drug combinations on gametocyte car‑ riage and gametocyte density along with asexual parasite density. Developing protocols of anti‑malarial drug efficacy that include gametocyte parameters related to both microscopic and submicroscopic gametocytaemias is important if drugs or drug combinations are to be strategically used in transmission‑blocking interventions in the context of malaria elimination. The present piece of opinion highlights the challenges in gametocyte detection and follow‑up and discuss the need for including the gametocyte parameter in anti‑malarial efficacy studies.

Background Giardia duodenalis is a protozoan parasite that can cause significant diarrhoeal diseases. Knowledge of population genetics is a prerequisite for ascertaining the invasion patterns of this parasite. In order to infer evolutionary patterns that could not be uncovered based on the morphological features, a population genetic study with the incorporation of molecular marker was carried out to access the genetic structure of G. duodenalis isolated from the Malaysian population and the global populations. Methods A total of 154 samples positive for Giardia, collected from different Malaysian communities, were subjected to DNA amplification and sequencing targeting three genetic loci (tpi, gdh, and bg). The tpi sequences together with sequences from the global data obtained from the NCBI GenBank were used for genetic diversity analyses including identification of haplotypes, haplotype diversity, nucleotide diversity, Tajima’s D and Fu and Li’s D, gene flow and genetic differentiation tests. Results Analysis of the Malaysian and global data showed that assemblages A, B, and E (the most prevalent assemblages in humans and animals), have different levels of genetic diversity. Assemblage B had the highest level of both haplotype diversity and nucleotide diversity, followed by assemblage E. The analysis also revealed population expansion and high gene flow in all assemblages. No clear genetic structure was observed across five continents (i.e., the Americas, Europe, Asia, Australia and Africa). However, median joining network of assemblage B formed a cluster that was exclusively isolated from Asia while other haplotypes were well dispersed across the continents. Conclusions This study provides new insight into the genetic diversity of Giardia assemblages in different geographical regions. The significant result shown by gene flow and genetic differentiation analyses as well as test of neutrality among the populations should have brought a clearer picture to the dynamics and distribution of Giardia infection.

Malaria is still a major public health problem in Yemen. More than 95% of the malaria cases are due to Plasmodium ‎falciparum‎. Recently in Yemen, the antimalarial treatment policy was changed from chloroquine (CQ) to artemisinin combination therapy (ACTs). However, CQ is still available and prescribed in the Yemeni market. The persistence of CQ resistance will be prolonged if the shift to ACT and the simultaneous withdrawal of CQ are not rigorously implemented. The aim of the current survey is to detect chloroquine-resistant mutations in P. falciparum chloroquine-resistance transporter (pfcrt) and P. falciparum multi-drug resistance-1 (pfmdr1) genes. These data will be important for future monitoring and assessment of antimalarial drug policy in Yemen. Blood specimens were collected from 735 individuals from different districts of the Hadhramout province, Yemen by house-to-house visit. Mutation-specific nested polymerase chain reaction (PCR) and restriction fragment length polymorphism (PCR–RFLP) methods were used to investigate the mutations in the pfmdr1(codons 86 and 1246) and pfcrt (codons 76, 271, 326, 356 and 371) genes. The overall prevalence of pfcrt mutations at codons 76, 271, 326 and 371 were 50.4%, 58.7%, 54.3% and 44.9%, respectively. All isolates had wild-type pfcrt 356 allele. The majority of pfmdr1 86 alleles (83.3%) and all pfmdr1 1246 alleles were wild type. There was no association between pfcrt mutations and symptomatology, gender and age groups. In conclusion, point mutations in codons 76, 271, 326 and 371 of pfcrt of P. falciparum are high suggesting a sustained high CQ resistance even after 4 years of shifting to ACTs. These findings warrant complete withdrawal of CQ use from the Yemeni market for P. falciparum and careful usage of CQ for treating Plasmodium vivax.

Introduction: Although toxoplasmosis is an important public health problem, there is scarcity of data on the disease available from Yemen. A cross-sectional survey was conducted in health facilities to determine seroprevalence of Toxoplasma gondii and associated risk factors among pregnant women in Sana’a, the capital city of Yemen. Methodology: A total of 593 pregnant women were included and examined for anti-T. gondii antibodies (Ab) using enzyme-linked immunosorbent assay. Bio and socio-demographic data were collected by pre-tested structured questionnaires through face-to-face interviews. Results: The overall seroprevalence of T. gondii was 45.4% (95% confidence interval: 41%–49%). The prevalence of anti-T. gondii IgG and IgM was 43.7 (95% CI: 40–%48%) and 9.1% (95% CI: 7%–12%), respectively. About 7.4 (95% CI: 6%–10%) of pregnant women were seropositive for both IgG and IgM Abs. Multivariate logistic regression analysis identified the following risk factors for toxplasmosis (IgG and/or IgM): age ≥ 25 years (adjusted OR: 2.02, 95% CI: 1.44–2.84, p < 0.001), rearing cats in the house (OR: 1.75, 95% CI: 1.20–2.55, p = 0.004), and contact with soil (OR: 1.90, 95% CI: 1.32–2.75, p = 0.001). Conclusions: The study reported high seroprevalence among pregnant women in Sana’a, Yemen, with a high proportion of pregnant women having a possibility of acute toxoplasmosis. This highlights the need for including routine screening for T. gondii in pregnant women in the country’s antenatal clinics. In addition, health education on the mode of transmission of toxoplasmosis should be provided for pregnant women in Yemen.

This study was carried out to investigate the prevalence and risk factors of Giardia infection among indigenous people in rural Malaysia. Faecal samples were collected from 1,330 participants from seven states of Malaysia and examined by wet mount and formalin-ether sedimentation methods while demographic, socioeconomic and environmental information was collected using a pre-tested questionnaire. The overall prevalence of Giardia infection was 11.6% and was significantly higher among those aged ≤ 12 years compared to their older counterparts. Multivariate logistic regression identified age of ≤12 years, lacking of toilet at household, not washing hands before eating, not washing hands after playing with animals, not boiling water before consumption, bathing in the river and not wearing shoes when outside as the significant risk factors of Giardia infection among these communities. Based on a multilocus genotyping approach (including tpi, gdh and bg gene sequences), 69 isolates were identified as assemblage A and 69 as assemblage B. No association between the assemblages and presence of symptoms was found. Providing proper sanitation, as well as provision of clean drinking water and proper health education regarding good personal hygiene practices will help significantly in reducing the prevalence and burden of Giardia infection in these communities.

Despite the intensive global efforts to control intestinal parasitic infections, the prevalence of soil-transmitted helminth (STH) infections is still very high in many developing countries particularly among children in rural areas. A randomized, double-blind, placebo-controlled trial was conducted on 250 Aboriginal schoolchildren in Malaysia to investigate the effects of a single high-dose of vitamin A supplementation (200 000 IU) on STH reinfection. The effect of the supplement was assessed at 3 and 6 months after receiving interventions; after a complete 3-day deworming course of 400 mg/daily of albendazole tablets. Almost all children (98.6%) were infected with at least one STH species. The overall prevalence of ascariasis, trichuriasis and hookworm infection was 67.8%, 95.5% and 13.4%, respectively. Reinfection rates of Ascaris, Trichuris and hookworm were high; at 6 months, assessment reached 80% of the prevalence reported before treatment. There were no significant differences in the reinfection rates and intensities of STH between vitamin A supplemented-children and those who received placebo at 3 and 6 months (p > 0.05). Vitamin A supplementation showed no protective effect against STH reinfection and this could be due to the high endemicity of STH in this community. Long-term interventions to reduce poverty will help significantly in reducing this continuing problem and there is no doubt that reducing intestinal parasitic infection would have a positive impact on the health, nutrition and education of these children. This trial was registered at clinicaltrials.gov as NCT00936091 .

We used a combined microscopy-molecular approach to determine the occurrence and identities of waterborne Giardia sp. cysts isolated from 18 separate, 10 l grab samples collected from a Malaysian zoo. Microscopy revealed that 17 of 18 samples were Giardia cyst positive with concentrations ranging from 1 to 120 cysts/l. Nine (52.9%) of the 17 cyst positive samples produced amplicons of which 7 (77.8%) could be sequenced. Giardia duodenalis assemblage A (6 of 7) and assemblage B (1 of 7), both infectious to humans, were identified at all sampling sites at the zoo. The presence of human infectious cysts raises public health issues, and their occurrence, abundance and sources should be investigated further. In this zoo setting, our data highlight the importance of incorporating environmental sampling (monitoring) in addition to routine faecal examinations to determine veterinary and public health risks, and water monitoring should be considered for inclusion as a separate element in hazard analysis, as it often has a historical (accumulative) connotation.

Ancylostoma ceylanicum is the only zoonotic hookworm species that is able to produce patent infections in humans with the majority of cases reported in South East Asia. Over the past few years, there have been an increasing number of studies investigating the prevalence of this parasitic zoonosis using molecular diagnostic tools and a single genetic locus as marker for species identification. As there can be limitations in using a single genetic locus for epidemiological studies and genetic discrimination, the complementary use of a more variable locus will provide additional evidence to support the zoonotic exchange of hookworm species between humans and animals. In the present study, the cytochrome c oxidase subunit 1 (cox 1) sequence of A. ceylanicum from positive human and animal fecal samples were determined and compared with published reference sequences. Phylogenetic analysis demonstrated that isolates of A. ceylanicum were divided into two clusters, one consisting 3 human isolates and the other comprising 19 isolates of human and animal origin from different geographical locations within Malaysia. The two groups of A. ceylanicum could be distinguished from one another through five fixed nucleotide differences at locations 891, 966, 1008, 1077 and 1083. The detection of genetically distinct groups and considerable level of genetic variation within the cox 1 sequence of A. ceylanicum might suggest potential haplotype-linked differences in zoonotic, epidemiological and pathobiological characteristics, a hypothesis that still needs further investigation.

BackgroundDespite evident success of malaria control in many sites in the Arabian Peninsula, malaria remains endemic in a few spots, in Yemen and south-west of Saudi Arabia. In addition to local transmission, imported malaria sustains an extra source of parasites that can challenge the strengths of local control strategies. This study examined the genetic diversity ofPlasmodium falciparumin Yemen and mutations of drug resistant genes, to elucidate parasite structure and distribution of drug resistance genotypes in the region.MethodsFive polymorphic loci (MSP-2,Pfg377and three microsatellites on chromosome 8) not involved in anti-malarial drug resistance, and four drug resistant genes (pfcrt,pfmdr1,dhfranddhps) were genotyped in 108P.falciparumisolates collected in three sites in Yemen: Dhamar, Hodeidah and Taiz.ResultsHigh diversity was seen in non-drug genes,pfg377(He = 0.66),msp-2(He = 0.80) and three microsatellites on chr 8, 7.7 kb (He = 0.88), 4.3 kb (He = 0.77) and 0.8 kb (He = 0.71). There was a high level of mixed-genotype infections (57%), with an average 1.8 genotypes per patient. No linkage disequilibrium was seen between drug resistant genes and the non-drug markers (p FSTvalues There was a high prevalence of mutations inpfmdr1,pfcrtanddhfr; with four mutantpfmdr1genotypes (NFCDD[57%], NFSND[21%], YFCDD[13%] and YFSND[8% ]), two mutantpfcrtgenotypes (CVIET[89%] and SVMNT[4%]) and one mutantdhfrgenotype (ICNI[53.7%]). However, nodhpsmutations were detected.ConclusionThe high diversity ofP.falciparumin Yemen is indicative of a large parasite reservoir, which represents a challenge to control efforts. The presence of two distinctpfcrtgenotype, CVIET and SVMNT, suggests that chloroquine resistance can possibly be related to a migratory path from Africa and Asia. The absence of the triple mutantdhfrgenotype (IRN) anddhpsmutations supports the use of artesunate + sulphadoxine-pyrimethamine as first-line therapy. However, the prevalentpfmdr1genotype NFSND [21%] has previously been associated with tolerance/resistance response to artemisinin combination therapy (ACT). Regular surveys are, therefore, important to monitor spread of pfmdr1 and dhfr mutations and response to ACT.

International audience; Background: The accuracy of the conclusions from in vivo efficacy anti-malarial drug trials depends on distinguishing between recrudescences and re-infections which is accomplished by genotyping genes coding P. falciparum merozoite surface 1 (MSP1) and MSP2. However, the reliability of the PCR analysis depends on the genetic markers’ allelic diversity and variant frequency. In this study the genetic diversity of the genes coding for MSP1 and MSP2 was obtained for P. falciparum parasites circulating in Yemen.Methods: Blood samples were collected from 511 patients with fever and screened for malaria parasites using Giemsa-stained blood films. A total 74 samples were infected with P. falciparum, and the genetic diversity was assessed by nested PCR targeting Pfmsp1 (Block2) and Pfmsp2 (block 3).Results: Overall, 58%, 28% and 54% of the isolates harboured parasites of the Pfmsp1 K1, MAD20 and RO33 allelic families, and 55% and 89% harboured those of the Pfmsp2 FC27 and 3D7 allelic families, respectively. For both genetic makers, the multiplicity of the infection (MOI) was significantly higher in the isolates from the foothills/coastland areas as compared to those from the highland (P

In Southeast Asia, Cryptosporidium and Giardia have been reported in countries such as Cambodia, Indonesia, Lao PDR, Malaysia, Myanmar, the Philippines, Singapore, Thailand and Vietnam. Some of the factors encouraging the transmission of Cryptosporidium and Giardia infections include rapid modernization, exponential population growth, greater human movement and the escalating numbers of HIV/AIDS individuals. The aim of this chapter is to consolidate available published reports on Cryptosporidium and Giardia in Southeast Asia and determine available information, fill up lacunae of knowledge and propose potential efforts that can lead to fruitful collaborations among scientists in the region in pursuit of a better understanding of these infections.

Giardia duodenalis is considered the most common intestinal parasite in humans worldwide. In Malaysia, many studies have been conducted on the epidemiology of giardiasis. However, there is a scarcity of information on the genetic diversity and the dynamics of transmission of G. duodenalis. The present study was conducted to identify G. duodenalis assemblages and sub-assemblages based on multilocus analysis of the glutamate dehydrogenase (gdh), beta-giardin (bg) and triose phosphate isomerase (tpi) genes. Faecal specimens were collected from 484 Orang Asli children with a mean age of 7 years and examined using light microscopy. Specimens positive for Giardia were subjected to PCR analysis of the three genes and subsequent sequencing in both directions. Sequences were edited and analysed by phylogenetic analysis. G. duodenalis was detected in 17% (84 of 484) of the examined specimens. Among them, 71 were successfully sequenced using at least one locus. Genotyping results showed that 30 (42%) of the isolates belonged to assemblage A, 32 (45%) belonged to assemblage B, while discordant genotype results were observed in 9 specimens. Mixed infections were detected in 43 specimens using a tpi-based assemblage specific protocol. At the sub-assemblages level, isolates belonged to assemblage A were AII. High nucleotide variation found in isolates of assemblage B made subtyping difficult to achieve. The finding of assemblage B and the anthroponotic genotype AII implicates human-to-human transmission as the most possible mode of transmission among Malaysian aborigines. The high polymorphism found in isolates of assemblage B warrants a more defining tool to discriminate assemblage B at the sub-assemblage level.

Visceral leishmaniasis (VL) is a debilitating, often fatal disease caused by Leishmania donovani complex; however, it is a neglected tropical disease. L. donovani complex comprises two closely related species, L. donovani that is mostly anthroponotic and L. infantum that is zoonotic. Differentiation between these two species is critical due to the differences in their epidemiology and pathology. However, they cannot be differentiated morphologically, and their speciation using isoenzyme-based methods poses a difficult task and may be unreliable. Molecular characterization is now the most reliable method to differentiate between them and to determine their phylogenetic relationships. The present study aims to characterize Leishmania species isolated from bone marrows of Yemeni pediatric patients using sequence analysis of the ribosomal internal transcribed spacer-1 (ITS1) gene. Out of 41 isolates from Giemsa-stained bone marrow smears, 25 isolates were successfully amplified by nested polymerase chain reaction and sequenced in both directions. Phylogenetic analysis using neighbor joining method placed all study isolates in one cluster with L. donovani complex (99% bootstrap). The analysis of ITS1 for microsatellite repeat numbers identified L. infantum in 11 isolates and L. donovani in 14 isolates. These data suggest the possibility of both anthroponotic and zoonotic transmission of VL-causing Leishmania species in Yemen. Exploring the possible animal reservoir hosts is therefore needed for effective control to be achieved.

Sashi S. Kommu1, Kamran Ahmed, Benjamin Challacombe, Prokar Dasgupta, Mohammed Shamim Khan, STILUS Academic Research Group (SARG) MRC Centre for Transplantation, King's College London, King's Health Partners, Department of Urology, Guy's Hospital Simulation and Interactive Learning (SaIL) Centre, Guy's & St Thomas NHS Foundation Trust Department of Urology, London, and STILUS Academic Research Group (SARG), London, UK

In the present study, 310 faecal samples from goats from eight different farms in Malaysia were tested for the presence of Giardia using a PCR-coupled approach. The nested PCR for SSU amplified products of the expected size (∼200 bp) from 21 of 310 (6.8%) samples. Sixteen of these 21 products could be sequenced successfully and represented six distinct sequence types. Phylogenetic analysis of the SSU sequence data using Bayesian Inference (BI) identified Giardia assemblages A, B and E. The identification of the ‘zoonotic’ assemblages A and B suggests that Giardia-infected goats represent a possible reservoir for human giardiasis in Malaysia.

Background Malaria is still a public health problem in Malaysia with chloroquine (CQ) being the first-line drug in the treatment policy of uncomplicated malaria. There is a scarcity in information about the magnitude of Plasmodium falciparum CQ resistance. This study aims to investigate the presence of single point mutations in the P. falciparum chloroquine-resistance transporter gene (pfcrt) at codons 76, 271, 326, 356 and 371 and in P. falciparum multi-drug resistance-1 gene (pfmdr1) at codons 86 and 1246, as molecular markers of CQ resistance. Methods A total of 75 P. falciparum blood samples were collected from different districts of Pahang state, Malaysia. Single nucleotide polymorphisms in pfcrt gene (codons 76, 271, 326, 356 and 371) and pfmdr1 gene (codons 86 and 1246) were analysed by using mutation-specific nested PCR and restriction fragment length polymorphism (PCR-RFLP) methods. Results Mutations of pfcrt K76T and pfcrt R371I were the most prevalent among pfcrt gene mutations reported by this study; 52% and 77%, respectively. Other codons of the pfcrt gene and the positions 86 and 1246 of the pfmdr1 gene were found mostly of wild type. Significant associations of pfcrt K76T, pfcrt N326S and pfcrt I356T mutations with parasitaemia were also reported. Conclusion The high existence of mutant pfcrt T76 may indicate the low susceptibility of P. falciparum isolates to CQ in Peninsular Malaysia. The findings of this study establish baseline data on the molecular markers of P. falciparum CQ resistance, which may help in the surveillance of drug resistance in Peninsular Malaysia.

Background Canine hookworm infection is endemic in Southeast Asian countries with a prevalence ranging from 70% to 100%, with zoonotic transmission representing a potentially significant public health concern. However, there are limited data available on the prevalence of canine hookworms in Malaysia. This study was conducted to determine the prevalence of hookworm and Ancylostoma species among dogs in Malaysia. Methods Faecal samples were collected from 221 dogs living in urban areas, rural areas and animal shelters in Selangor. Faecal samples were processed using the formal-ether concentration technique followed by wet mount preparation and iodine staining for the detection of hookworm eggs. Samples positive for hookworm eggs were examined using PCR, targeting ITS2 and 28 s rRNA region, and subsequently sequenced in both directions. The sequences were phylogenetically analysed using MrBayes for Bayesian Inference. Results The overall prevalence of hookworm among dogs was 48% (95%CI; 41.41–54.95). Rural stray dogs had the highest prevalence 71.4% (95%CI; 61.13–81.49) followed by urban stray dogs, recording 48% (95%CI; 34.15–61.85) and lastly dogs in shelters with 28.7% (95%CI; 19.56–37.84). Logistic regression identified rural stray dogs as a high risk group (OR = 4.55, 95%; 2.50–8.31) and keeping dogs in shelters as a protective factor (OR = 0.24, 95%; 0.14–0.43). Molecular methods identified both Ancylostoma ceylanicum and Ancylostoma caninum with A. ceylanicum being predominant among urban stray dogs. Rural dogs had a higher prevalence of A. caninum than A. ceylanicum, while both species showed equal distribution among dogs in shelters. Phylogenetic analysis placed A. ceylanicum isolated from dogs in one group with A. ceylanicum human isolates. Conclusion This study indicates that dogs have the potential to act as reservoir hosts of human hookworm infection in Malaysia. This finding necessitates the inclusion of dogs in any interventions to combat hookworm in the country.

Although intestinal parasitic infections (IPI) among children remain a global issue, the current information on such infections in Albanian children is very limited. A cross-sectional study of the IPI in 321 children living in the Albanian counties of Tirana (152) and Elbasan (169) was therefore conducted in 2008, with a pre-tested standard questionnaire employed to gather the relevant personal and clinical data. Using formalin–ether concentration and permanent stains, stool samples were examined microscopically for the ova, cysts and oocysts of any parasites. The overall prevalence of IPI was 19% (61 of 321), with protozoan infections (11·5%) apparently more common than infections with soil-transmitted helminths (STH; 8·1%). Giardia duodenalis was the parasite most frequently detected (10·9%), followed by hookworm (5·6%), Ascaris lumbricoides (1·9%), Trichuris trichiura (0·6%), Cryptosporidium (0·3%) and Entamoeba histolytica/dispar (0·3%). The results of a univariate analysis indicated that the children from Tirana county were significantly more likely to be found infected with STH compared with the children from Elbasan county (12·5% v. 4·1%; P = 0·006). Children sampled in the community were also more likely to be found STH-positive than the children sampled as they attended hospitals and health clinics (10·5% v. 6·0%) but this difference did not reach statistical significance. The children found STH-positive were five times more likely to be suffering from diarrhoea than the other children checked in clinical settings (P = 0·004) and were also more likely to be suffering from abdominal pain (P = 0·054) and/or diminished appetite (P = 0·016).

title/Between June 2008 and March 2009, a cross-sectional study of human malaria was carried out in four governorates of Yemen, two (Taiz and Hodiedah) representing the country's highlands and the others (Dhamar and Raymah) the country's coastal plains/foothills. The main aims were to determine the prevalences of Plasmodium infection among 455 febrile patients presenting for care at participating health facilities and to investigate the potential risk factors for such infection. Malarial infection was detected in 78 (17·1%) of the investigated patients and was more likely to be detected among the febrile patients from the highlands than among those presenting in the coastal plains/foothills (22·6% v.13·9%; χ(2)=10·102; P=0·018). Binary logistic-regression models identified low household income [odds ratio (OR)=13·52; 95% confidence interval (CI)=2·62-69·67; P=0·002], living in a household with access to a water pump (OR=4·18; CI=1·60-10·96; P=0·004) and living in a household near a stream (OR=4·43; CI=1·35-14·56; P=0·014) as significant risk factors for malarial infection in the highlands. Low household income was the only significant risk factor identified for such infection in the coastal plains and foothills (OR = 8·20; CI=1·80-37·45; P=0·007). It is unclear why febrile patients in the highlands of Yemen are much more likely to be found to have malarial infection than their counterparts from the coastal plains and foothills. Although it is possible that malarial transmission is relatively intense in the highlands, it seems more likely that, compared with those who live at lower altitudes, those who live in the highlands are less immune to malaria, and therefore more likely to develop febrile illness following malarial infection. Whatever the cause of the symptomatic malarial infection commonly found in the highlands of Yemen, it is a matter of serious concern that should be addressed in the national strategy to control malaria.

Cryptosporidium is a coccidian parasite that is prevalent worldwide, some species of which cause morbidity in both immunocompromised and immunocompetent individuals. The prevalence and predictors of Cryptosporidium infection, and its effect on nutritional status, have recently been explored among 276 children (141 boys and 135 girls, aged 2-15 years) in aboriginal (Orang Asli) villages in the Malaysian state of Selangor. Faecal smears were examined by the modified Ziehl-Neelsen staining technique while socio-economic data were collected using a standardized questionnaire. Nutritional status was assessed by anthropometric measurements. Cryptosporidium infection, which was detected in 7.2% of the aboriginal children, was found to be significantly associated with low birthweight (≤2.5 kg), being part of a large household (with more than seven members) and prolonged breast feeding (>2 years). The output of a binary logistic regression confirmed that large household size was a significant predictor of Cryptosporidium infection (giving an odds ratio of 2.15, with a 95% confidence interval of 1.25-5.02). Cryptosporidium infection is clearly a public-health problem among the aboriginal children of Selangor, with person-to-person the most likely mode of transmission.

BACKGROUND: Intestinal protozoan diseases in Yemen are a significant health problem with prevalence ranging from 18% to 27%. The present study is a cross-sectional study aimed at determining the factors associated with the high prevalence of intestinal protozoan infections among patients seeking health care in Sana'a City, the capital of Yemen. METHODOLOGY/PRINCIPAL FINDINGS: Stool samples were collected from 503 patients aged between 1 and 80 years old; 219 were males and 284 females. Biodata were collected via pretested standard questionnaire. Faecal samples were processed and examined for (oo)cysts or ova using a wet mount preparation after formal-ether concentration technique. Cryptosporidium oocysts were detected using the Ziehl-Neelsen staining technique. The overall prevalence of intestinal protozoan infections was 30.9%. Infection rates of Giardia duodenalis, Entamoeba histolytica/dispar and Cryptosporidium were 17.7%, 17.1% and 1%, respectively. Other parasites detected included Ascaris lumbricoides (2.4%), Schistosoma mansoni (0.3%), Hymenolepis nana (1.4%) and Enterobius vermicularis (0.4%). Multivariate analysis using forward stepwise logistic regression based on intestinal protozoan infections showed that contact with animals (OR = 1.748, 95% CI = 1.168-2.617) and taking bath less than twice a week (OR = 1.820, 95% CI = 1.192-2.779) were significant risk factors of protozoan infections. CONCLUSIONS/SIGNIFICANCE: This present study indicated that intestinal protozoan infections are still a public health problem in Yemen, with Giardia and Entamoeba infections being most common. Statistical analysis indicated that low personal hygiene and contact with animals were important predictors for intestinal protozoan infections. As highlighted in this study, in order to effectively reduce these infections, a multi-sectoral effort is needed. Preventive measures should include good hygienic practices, good animal husbandry practices, heightened provision of educational health programs, health services in all governorates including rural areas. Furthermore, it is also essential to find radical solutions to the recent water crises in Yemen.

Nine 50-l surface water samples from a Malaysian recreational lake were examined microscopically using an immunomagnetisable separation-immunofluorescent method. No Cryptosporidium oocysts were detected, but 77.8% of samples contained low numbers of Giardia cysts (range, 0.17-1.1 cysts/l), which were genetically characterised by SSU rRNA gene sequencing. Genotype analyses indicated the presence of Giardia duodenalis assemblage A suggesting potential risk to public health. The present study represents the first contribution to our knowledge of G. duodenalis assemblages in Malaysian recreational water.

We present a microfluidic bioreactor for culturing high-density arrays of hepatocytes in a tissue-like micro-architecture. The microfluidic environment mimicked physiological liver mass transport, enabling sustained culture of high density cells (>2,000 cells/mm(2)) without nutrient limitation for over 1 week. The key feature of this design was a microporous microfluidic barrier that formed a sieved-pocket to concentrate cells during loading. Nutrient depletion within the cell mass was avoided by maintaining a continuous flow of medium (10 microl/day) that diffused across the porous barrier. Human hepatoma cells (HepG2/C3A) remained viable and functional as demonstrated by fluorescent viability assays and secretion of albumin for the one-week culture period.

The mechanism by which exposure to the industrial solvent benzene (BZ) can result in aplastic anemia or leukemia (see Snyder and Kalf, 1994 for review) has been under intensive investigation. Topoisomerase (topo) II inhibitors are used clinically as cancer chemotherapeutic agents and can also lead to secondary AML (see Felix, 1998; Leone et al., 1999 for review). Hydroquinone (HQ), a hematotoxic metabolite of BZ, has been shown to inhibit purified human topo II (Hutt and Kalf, 1996; Frantz et al., 1996). It has therefore been suggested that some of the hematotoxic effects of BZ may be mediated by the ability of some of its metabolites to inhibit topo II activity (Whysner, 2000). In the studies presented here a normal, IL-3 dependent mouse myeloblast cell line (32D.3 (G)) was used to determine whether the hematotoxic effects of BZ may result from inhibition of topo II by HQ. The 32D.3 (G) myeloblasts undergo differentiation and maturation to neutrophils upon treatment with the physiological inducer, granulocyte colony-stimulating factor (G-CSF) (Metcalf, 1985; Valtieri et al., 1987). In this system HQ treatment inhibits apoptosis, induces differentiation of myeloblasts to the myelocyte stage, and blocks maturation to the neutrophil (Hazel et al., 1995), a series of events similar to those observed in AML, which is characterized by clonal expansion of immature granulocytes (Lowenberg and Delwel, 1991). There is evidence linking topo II activity to differentiation and maturation in the HL-60, human leukemia cell line. In this system 3-nitrobenzothiazolo[3,2-a]quinolinium, an inhibitor of topo II activity, has been shown to induce incomplete myeloid differentiation (Baez and Sepulveda, 1992).

Transient transfection experiments in which three different estrogen response element-containing reporter genes were cotransfected into HeLa cells, together with constitutively expressed estrogen receptor (ER) constructs, demonstrate that activation of the transcription of the reporter genes by epidermal growth factor (EGF) and by cholera toxin with 3-isobutyl-1-methyl-xanthine, which elevate cellular cAMP, is dependent upon the presence of functional ER. Cotransfection of the reporter genes with truncated versions of the ER shows that the two non-ligand activators of ER require different regions of the receptor to produce their effects on transcription. EGF acts primarily by means of transactivation domain AF-1, whereas cAMP acts via transactivation domain AF-2 of the ER. A point mutation that removes a major site of inducible phosphorylation within the AF-1 domain of the ER abolishes the response to EGF, but the response to estradiol and cAMP is retained. Specific inhibition of cAMP-activated protein kinase (protein kinase A) prevents the response to elevated cAMP but does not affect EGF or estradiol responses. Overexpression of the protein kinase A catalytic subunit in HeLa cells results in an amplified response to estradiol, similar to that induced by cholera toxin with 3-isobutyl-1-methyl-xanthine. Comparable experiments performed using COS-1 cells produce similar results but also reveal cell type- and promoter-specific aspects of the activation mechanisms. Apparently, the ER may be activated by three different signal molecules, estradiol, EGF, and cAMP, each using a mechanism that is distinguishable from that of the others.

The response of two endogenous, estrogen-induced genes, LIV-1 and pS2, to growth factor stimulation of MCF-7 cells was examined. Epidermal growth factor (EGF), transforming growth factor alpha (TGF alpha) and insulin-like growth factor-1 (IGF-1) were each able to induce an increase in the two mRNAs in the absence of estradiol, and their effects were additive to that of an optimally inducing concentration (10(-8) M) of the hormone. Induction by EGF and TGF alpha, but not by IGF-1, were also additive to induction by a saturating concentration (2 microg/ml) of insulin. TGFbeta, an antimitogenic growth factor for MCF-7 cells, did not induce LIV-1 or pS2 mRNA but inhibited induction by estradiol. Increases in mRNA were shown to reflect increases in specific gene transcription. Induction by growth factors, but not by estradiol, was dependent upon protein synthesis. Induction by both growth factors and estradiol was inhibited by the pure antiestrogen, ICI 164384 (ICI), and by the mixed agonist/antagonist, tamoxifen. Despite differences in patterns of expression in vivo and in vitro, both LIV-1 and pS2 appeared to be responsive to growth factors via a mechanism distinct from that of estradiol but requiring the estrogen receptor.

Background Schistosomiasis, one of the most prevalent neglected tropical diseases, is a life-threatening public health problem in Yemen especially in rural communities. This cross-sectional study aims to determine the prevalence and associated risk factors of schistosomiasis among children in rural Yemen. Methods/Findings Urine and faecal samples were collected from 400 children. Urine samples were examined using filtration technique for the presence of Schistosoma haematobium eggs while faecal samples were examined using formalin-ether concentration and Kato Katz techniques for the presence of S. mansoni. Demographic, socioeconomic and environmental information were collected via a validated questionnaire. Overall, 31.8% of the participants were found to be positive for schistosomiasis; 23.8% were infected with S. haematobium and 9.3% were infected with S. mansoni. Moreover, 39.5% of the participants were anaemic whereas 9.5% had hepatosplenomegaly. The prevalence of schistosomiasis was significantly higher among children aged >10 years compared to those aged ≤10 years (P, Author Summary Schistosomiasis remains one of the most serious and prevalent diseases worldwide. Despite intensive control efforts by the government and international bodies, schistosomiasis is the second cause of death, after malaria, in Yemen, with an estimated 3 million cases. We screened 400 children in rural areas of five provinces in Yemen for the presence of schistosomiasis. Overall, 31.8% of the children were found to be positive for schistosomiasis; 23.8% were infected by Schistosoma haematobium, and 9.3% were infected by S. mansoni. The study identified the presence of other family members infected with schistosomiasis, low household monthly income, using unsafe water supply as a source for drinking water, living nearby stream/spring and/or pool/pond as the key factors significantly associated with schistosomiasis in these communities. Innovative and integrated control measures to control this infection should be implemented among this population. Periodic school-based and community-based drug distribution, health education, provision of clean and safe drinking water, introduction of proper sanitation will help to reduce the prevalence and morbidity of schistosomiasis among these communities.

The level of oestrogen-responsive gene expression in breast tumours has been proposed as a predictor of the response of the tumour to endocrine (anti-oestrogen) therapy. We demonstrate that different oestrogen-responsive genes may differ in their responses to other hormones. pLIV-1 and pS2 are two oestrogen-regulated genes that are expressed in the MCF-7 human breast cancer cell line. We show that pLIV-1 mRNA, but not pS2 mRNA, is also induced, to a lesser extent, by progesterone, 5 alpha-dihydrotestosterone and dexamethasone. For pLIV-1, combinations of these hormones with oestradiol and with the pure anti-oestrogen, ICI 164384, indicate that the mechanism of its response to these other steroid hormones is clearly separable from its response to oestrogen. Such behaviour in breast tumours in vivo could explain the lack of absolute correlation between marker gene expression and anti-oestrogen sensitivity and between the expression of individual marker genes.

Background Malaria is an endemic disease in Yemen and is responsible for 4.9 deaths per 100,000 population per year and 43,000 disability adjusted life years lost. Although malaria in Yemen is caused mainly by Plasmodium falciparum and Plasmodium vivax, there are no sequence data available on the two species. This study was conducted to investigate the distribution of the Plasmodium species based on the molecular detection and to study the molecular phylogeny of these parasites. Methods Blood samples from 511 febrile patients were collected and a partial region of the 18 s ribosomal RNA (18 s rRNA) gene was amplified using nested PCR. From the 86 positive blood samples, 13 Plasmodium falciparum and 4 Plasmodium vivax were selected and underwent cloning and, subsequently, sequencing and the sequences were subjected to phylogenetic analysis using the neighbor-joining and maximum parsimony methods. Results Malaria was detected by PCR in 86 samples (16.8%). The majority of the single infections were caused by P. falciparum (80.3%), followed by P. vivax (5.8%). Mixed infection rates of P. falciparum + P. vivax and P. falciparum + P. malariae were 11.6% and 2.3%, respectively. All P. falciparum isolates were grouped with the strain 3D7, while P. vivax isolates were grouped with the strain Salvador1. Phylogenetic trees based on 18 s rRNA placed the P. falciparum isolates into three sub-clusters and P. vivax into one cluster. Sequence alignment analysis showed 5-14.8% SNP in the partial sequences of the 18 s rRNA of P. falciparum. Conclusions Although P. falciparum is predominant, P. vivax, P. malariae and mixed infections are more prevalent than has been revealed by microscopy. This overlooked distribution should be considered by malaria control strategy makers. The genetic polymorphisms warrant further investigation.

Immunohistochemical staining of formalin fixed, paraffin embedded tissue sections of OSF for MMPs-1,2,9 and their tissue inhibitors TIMP-1and 2 was performed using monospecific antibodies coupled with gelatin zymography (MMP-2 and 9) for measuring enzymatic activity quantitatively and for distinguishing the active from the inactive variants of enzymes. The present study, contrary to earlier reports, recorded statistically significant increase in the levels of stromal expression of MMP-1, MMP-2 and MMP-9 and TIMP-1 and TIMP-2 using monospecific antibodies reacting against tissue antigens. The simultaneous increase in reactivity of MMPs and TIMPs poise difficulty in interpretingthe results of this study. The possible reasons for this result, against the backdrop of existing knowledge, were attempted in this study.

Six UV induced cycloheximide-sensitive revertants were isolated from the cyh1-C7 strain of Schizosaccharomyces pombe which is resistant to cycloheximide. In all cases reversion to sensitivity was due to a forward mutation in a second suppressor gene. Genetical analysis showed that at least two genes, designated scr1 and scr2 (scr=suppression of cycloheximide resistance) were involved. Both scr1 and scr2 suppressed the resistance of six independently isolated alleles at the cyh1 locus. They had no effect on two known nonsense mutations in the ade7 locus. The cyh1-C7 strain has an altered 60S ribosomal protein which can be detected by two-dimensional polyacrylamide gel electrophoresis. In two suppressed strains, cyh1-C7 scr1 and cyh1-C7 scr2, the original altered protein was present. However no further ribosomal protein differences were observed which could be correlated with the presence of the scr genes. Both scr mutations conferred cold sensitivity on the organism indicating that they were of the missense type. Hence it seems certain that scr1 and scr2 are not mutations in tRNA genes leading to either nonsense or missense suppression. There is however no direct evidence that they code for ribosomal proteins and exert their effect on cyh1-C7 at the ribosomal level.