Prion Protein Polymorphisms and Estimation of Risk of Scrapie in East Asian Sheep

Allele and genotype frequency distributions of prion protein (PrP) polymorphisms at three codons, 136, 154, and 171, in East Asian sheep were determined by PCR–RFLP analysis using 553 animals from nine local breeds of the northern group and four local breeds of the southern group. Based on the genotype distribution, the risk score for scrapie was estimated. Among the local breeds, ARQ appeared predominantly (0.7701–1), followed by ARH and ARR. From such a biased allele distribution, it was difficult to ascertain the prevalent genetic relationships. A marked difference in allele frequencies between the northern and southern groups was seen (P < 0.0001). The East Asian sheep had ARQ at the highest frequency (0.8834); in European sheep it was 0.5317. According to an assessment of scrapie risk in the PrP genotype classes, the predominant ARQ/ARQ in East Asian sheep corresponded to the risk score of R4. This finding suggests that East Asian sheep have a high level of genetic susceptibility to scrapie.     

Phylogenetic Relationships among Indigenous Sheep Populations in East Asia based on Five Informative Blood Protein and Nonprotein Polymorphisms

The phylogenetic relationships among 23 local sheep breeds and varieties in East Asia were determined based on allele frequency data for five informative and polymorphic blood protein and nonprotein loci (transferrin, arylesterase, hemoglobin-β, X-protein, and potassium transport) using electrophoresis and ion-densitometric techniques. Comparatively striking differences in the frequency distributions, especially of hemoglobin-β and X-protein alleles, were seen between the northern population of the Khalkhas, Bhyanglung, Baruwal, Jakar, Sakten, and five Chinese breeds and the southern population of the Bengal, Kagi, Lampuchhre, Myanmar, and Sipsu breeds. Clustering analyses using UPGMA and NJ methods and principal component analysis, using the data of all populations and loci tested, clearly demonstrated the difference. The two population groups are divided by the boundary of the Himalayas, and each diverged into three subgroups: Mongolian, Tibetan, and Himalayan groups in the north and Indian groups I, II, and III in the south. It was noted that the genetic differentiation of populations was more distinct in the northern group. These findings strongly suggest the existence of at least two large and phylogenetically different gene pools of sheep in East Asia.     

Additional materials of Myanmarpithecus yarshensis (Amphipithecidae, Primates) from the middle Eocene Pondaung Formation

Myanmarpithecus yarshensis is an amphipithecid primate from the middle Eocene Pondaung Formation in Myanmar. It was previously known based on maxillary fragments with P⁴–M³ and mandibular fragments with C–P₃ and M_2–3. This study reports new materials for the genus, including a humeral head fragment, a lingual fragment of the right M², a lingual fragment of the right M³, and a left I¹. These new materials were collected from approximately the same point, and likely belonged to the same individual. The upper molar morphology and size of the new materials show similarity to those of the type specimen, indicating that the new materials can be assigned to M. yarshensis. The humeral head is the first postcranial element that is associated with dental materials for amphipithecids. The morphological similarity between the previously reported larger humerus and this specimen confirms the assignment of the former specimen to Amphipithecidae and suggests common locomotor adaptations in the family. The upper central incisor is large relative to the molar fragments, but is within the variation among extant platyrrhines. The tooth is spatulate-shaped and high crowned, and lacks the mesial process, indicating similarity to I¹ of haplorhines and clear differences from that of adapoids. It has been suggested that amphipithecids, including Myanmarpithecus, have affinities with notharctine adapoids, but the morphology of I¹ does not support the notharctine hypothesis of the Amphipithecidae.     

Malaria Hotspots Drive Hypoendemic Transmission in the Chittagong Hill Districts of Bangladesh

Background

Malaria is endemic in 13 of 64 districts of Bangladesh, representing a population at risk of about 27 million people. The highest rates of malaria in Bangladesh occur in the Chittagong Hill Districts, and Plasmodium falciparum (predominately chloroquine resistant) is the most prevalent species.

Methods

The objective of this research was to describe the epidemiology of symptomatic P. falciparum malaria in an area of Bangladesh following the introduction of a national malaria control program. We carried out surveillance for symptomatic malaria due to P. falciparum in two demographically defined unions of the Chittagong Hill Districts in Bangladesh, bordering western Myanmar, between October 2009 and May 2012. The association between sociodemographics and temporal and climate factors with symptomatic P. falciparum infection over two years of surveillance data was assessed. Risk factors for infection were determined using a multivariate regression model.

Results

472 cases of symptomatic P. falciparum malaria cases were identified among 23,372 residents during the study period. Greater than 85% of cases occurred during the rainy season from May to October, and cases were highly clustered geographically within these two unions with more than 80% of infections occurring in areas that contain approximately one-third of the total population. Risk factors statistically associated with infection in a multivariate logistic regression model were living in the areas of high incidence, young age, and having an occupation including jhum cultivation and/or daily labor. Use of long lasting insecticide-treated bed nets was high (89.3%), but its use was not associated with decreased incidence of infection.

Conclusion

Here we show that P. falciparum malaria continues to be hypoendemic in the Chittagong Hill Districts of Bangladesh, is highly seasonal, and is much more common in certain geographically limited hot spots and among certain occupations.     

Role of the Vibrio cholerae Matrix Protein Bap1 in Cross-Resistance to Antimicrobial Peptides

Outer membrane vesicles (OMVs) that are released from Gram-negative pathogenic bacteria can serve as vehicles for the translocation of effectors involved in infectious processes. In this study we have investigated the role of OMVs of the Vibrio cholerae O1 El Tor A1552 strain in resistance to antimicrobial peptides (AMPs). To assess this potential role, we grew V. cholerae with sub-lethal concentrations of Polymyxin B (PmB) or the AMP LL-37 and analyzed the OMVs produced and their effects on AMP resistance. Our results show that growing V. cholerae in the presence of AMPs modifies the protein content of the OMVs. In the presence of PmB, bacteria release OMVs that are larger in size and contain a biofilm-associated extracellular matrix protein (Bap1). We demonstrated that Bap1 binds to the OmpT porin on the OMVs through the LDV domain of OmpT. In addition, OMVs from cultures incubated in presence of PmB also provide better protection for V. cholerae against LL-37 compared to OMVs from V. cholerae cultures grown without AMPs or in presence of LL-37. Using a bap1 mutant we showed that cross-resistance between PmB and LL-37 involved the Bap1 protein, whereby Bap1 on OMVs traps LL-37 with no subsequent degradation of the AMP.     

Staphylococcus aureus α-Toxin-Dependent Induction of Host Cell Death by Membrane-Derived Vesicles

Staphylococcus aureus causes a wide spectrum of infections in humans, ranging from superficial cutaneous infections, infections in the circum-oral region, to life-threatening bacteremia. It was recently demonstrated that Gram-positive organisms such as S. aureus liberate membrane-derived vesicles (MVs), which analogously to outer membrane vesicles (OMVs) of Gram-negative bacteria can play a role in delivering virulence factors to host cells. In the present study we have shown that cholesterol-dependent fusion of S. aureus MVs with the plasma membrane represents a route for delivery of a key virulence factor, α-toxin (α-hemolysin; Hla) to human cells. Most S. aureus strains produce this 33-kDa pore-forming protein, which can lyse a wide range of human cells, and induce apoptosis in T-lymphocytes. Our results revealed a tight association of biologically active α-toxin with membrane-derived vesicles isolated from S. aureus strain 8325-4. Concomitantly, α-toxin contributed to HeLa cell cytotoxicity of MVs, and was the main vesicle-associated protein responsible for erythrocyte lysis. In contrast, MVs obtained from an isogenic hla mutant were significantly attenuated with regards to both causing lysis of erythrocytes and death of HeLa cells. This is to our knowledge the first recognition of an S. aureus MV-associated factor contributing to host cell cytotoxicity.     

Is Plasmodium vivax Malaria a Severe Malaria?: A Systematic Review and Meta-Analysis

Background

Plasmodium vivax is one of the major species of malaria infecting humans. Although emphasis on P. falciparum is appropriate, the burden of vivax malaria should be given due attention. This study aimed to synthesize the evidence on severe malaria in P. vivax infection compared with that in P. falciparum infection.

Methods/Principal Findings

We searched relevant studies in electronic databases. The main outcomes required for inclusion in the review were mortality, severe malaria (SM) and severe anaemia (SA). The methodological quality of the included studies was assessed using the Newcastle-Ottawa Scale. Overall, 26 studies were included. The main meta-analysis was restricted to the high quality studies. Eight studies (n = 27490) compared the incidence of SM between P. vivax infection and P. falciparum mono-infection; a comparable incidence was found in infants (OR: 0.45, 95% CI:0.04–5.68, I ²:98%), under 5 year age group (OR: 2.06, 95% CI: 0.83–5.1, I ²:83%), the 5–15 year-age group (OR: 0.6, 95% CI: 0.31–1.16, I ²:81%) and adults (OR: 0.83, 95% CI: 0.67–1.03, I ²:25%). Six studies reported the incidences of SA in P. vivax infection and P. falciparum mono-infection; a comparable incidence of SA was found among infants (OR: 3.47, 95%:0.64–18.94, I ²: 92%), the 5–15 year-age group (OR:0.71, 95% CI: 0.06–8.57, I ²:82%). This was significantly lower in adults (OR:0.75, 95% CI: 0.62–0.92, I ²:0%). Five studies (n = 71079) compared the mortality rate between vivax malaria and falciparum malaria. A lower rate of mortality was found in infants with vivax malaria (OR:0.61, 95% CI:0.5–0.76, I ²:0%), while this was comparable in the 5–15 year- age group (OR: 0.43, 95% CI:0.06–2.91, I ²:84%) and the children of unspecified-age group (OR: 0.77, 95% CI:0.59–1.01, I ²:0%).

Conclusion

Overall, the present analysis identified that the incidence of SM in patients infected with P. vivax was considerable, indicating that P. vivax is a major cause of SM. Awareness of the clinical manifestations of vivax malaria should prompt early detection. Subsequent treatment and monitoring of complications can be life-saving.     

Characterization of dominantly negative mutant ClyA cytotoxin proteins in Escherichia coli

We report studies of the subcellular localization of the ClyA cytotoxic protein and of mutations causing defective translocation to the periplasm in Escherichia coli. The ability of ClyA to translocate to the periplasm was abolished in deletion mutants lacking the last 23 or 11 amino acid residues of the C-terminal region. A naturally occurring ClyA variant lacking four residues (183 to 186) in a hydrophobic subdomain was retained mainly in the cytosolic fraction. These mutant proteins displayed an inhibiting effect on the expression of the hemolytic phenotype of wild-type ClyA. Studies in vitro with purified mutant ClyA proteins revealed that they were defective in formation of pore assemblies and that their activity in hemolysis assays and in single-channel conductance tests was at least 10-fold lower than that of the wild-type ClyA. Tests with combinations of the purified proteins indicated that mutant and wild-type ClyA interacted and that formation of heteromeric assemblies affected the pore-forming activity of the wild-type protein. The observed protein-protein interactions were consistent with, and provided a molecular explanation for, the dominant negative feature of the mutant ClyA variants.     

Assessment of secondary zinc reserves of nations

When  addressing the sustainable use of metals, one must consider not only primary metals in the natural environment but also alternative resources, such as secondary metals found in society. For that purpose, elucidating the availability of secondary metals, that is, secondary metal reserves, is important. A classification framework of the secondary resources was applied to investigate its applicability to zinc and to assess the secondary zinc reserves and resources of major targeted countries. Our estimates show that Japan and the United States have secondary zinc reserves of 14 and 13 Mt, respectively, and showed the total estimated amount of secondary zinc reserves of the study countries is equivalent to about 24% of the global primary zinc reserves. On a per‐capita basis, France, Germany, and Japan have the largest secondary zinc reserves. The application of a classification framework showed that a considerable amount of secondary zinc resources is found in landfills, providing a future potential target for secondary zinc landfill mining. The framework provides details about the sizes and locations of secondary zinc resources. This information is useful for both industry and policy makers to maximize access to valuable secondary zinc sources. This study also highlights the necessity for the integrated management of primaszzry and secondary zinc resources.     

Geographical distribution and genetic diversity of Plasmodium vivax reticulocyte binding protein 1a correlates with patient antigenicity

Plasmodium vivax is the most widespread cause of human malaria. Recent reports of drug resistant vivax malaria and the challenge of eradicating the dormant liver forms increase the importance of vaccine development against this relapsing disease. P. vivax reticulocyte binding protein 1a (PvRBP1a) is a potential vaccine candidate, which is involved in red cell tropism, a crucial step in the merozoite invasion of host reticulocytes. As part of the initial evaluation of the PvRBP1a vaccine candidate, we investigated its genetic diversity and antigenicity using geographically diverse clinical isolates. We analysed pvrbp1a genetic polymorphisms using 202 vivax clinical isolates from six countries. Pvrbp1a was separated into six regions based on specific domain features, sequence conserved/polymorphic regions, and the reticulocyte binding like (RBL) domains. In the fragmented gene sequence analysis, PvRBP1a region II (RII) and RIII (head and tail structure homolog, 152–625 aa.) showed extensive polymorphism caused by random point mutations. The haplotype network of these polymorphic regions was classified into three clusters that converged to independent populations. Antigenicity screening was performed using recombinant proteins PvRBP1a-N (157–560 aa.) and PvRBP1a-C (606–962 aa.), which contained head and tail structure region and sequence conserved region, respectively. Sensitivity against PvRBP1a-N (46.7%) was higher than PvRBP1a-C (17.8%). PvRBP1a-N was reported as a reticulocyte binding domain and this study identified a linear epitope with moderate antigenicity, thus an attractive domain for merozoite invasion-blocking vaccine development. However, our study highlights that a global PvRBP1a-based vaccine design needs to overcome several difficulties due to three distinct genotypes and low antigenicity levels.