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Malaria remains one of the most prevalent and lethal human infectious diseases worldwide. A comprehensive characterization of antibody responses to blood stage malaria is essential to support the development of future vaccines, sero-diagnostic tests, and sero-surveillance methods. We constructed a proteome array containing 4441 recombinant proteins expressed by the blood stages of the two most common human malaria parasites, P. falciparum (Pf) and P. vivax (Pv), and used this array to screen sera of Papua New Guinea children infected with Pf, Pv, or both (Pf/Pv) that were either symptomatic (febrile), or asymptomatic but had parasitemia detectable via microscopy or PCR. We hypothesized that asymptomatic children would develop antigen-specific antibody profiles associated with antidisease immunity, as compared with symptomatic children. The sera from these children recognized hundreds of the arrayed recombinant Pf and Pv proteins. In general, responses in asymptomatic children were highest in those with high parasitemia, suggesting that antibody levels are associated with parasite burden. In contrast, symptomatic children carried fewer antibodies than asymptomatic children with infections detectable by microscopy, particularly in Pv and Pf/Pv groups, suggesting that antibody production may be impaired during symptomatic infections. We used machine-learning algorithms to investigate the relationship between antibody responses and symptoms, and we identified antibody responses to sets of Plasmodium proteins that could predict clinical status of the donors. Several of these antibody responses were identified by multiple comparisons, including those against members of the serine enriched repeat antigen family and merozoite protein 4. Interestingly, both P. falciparum serine enriched repeat antigen-5 and merozoite protein 4 have been previously investigated for use in vaccines. This machine learning approach, never previously applied to proteome arrays, can be used to generate a list of potential seroprotective and/or diagnostic antigens candidates that can be further evaluated in longitudinal studies.

Extreme diversity of the major Plasmodium falciparum antigen, PfEMP1, poses a barrier to identifying targets of immunity to malaria. Here, we used protein microarrays containing hundreds of variants of the DBLα domain of PfEMP1 to cover the diversity of Papua New Guinean (PNG) parasites. Probing the plasma of a longitudinal cohort of malaria-exposed PNG children showed that group 2 DBLα antibodies were moderately associated with a lower risk of uncomplicated malaria, whereas individual variants were only weakly associated with clinical immunity. In contrast, antibodies to 85 individual group 1 and 2 DBLα variants were associated with a 70%-100% reduction in severe malaria. Of these, 17 variants were strong predictors of severe malaria. Analysis of full-length PfEMP1 sequences from PNG samples shows that these 17 variants are linked to pathogenic CIDR domains. This suggests that whereas immunity to uncomplicated malaria requires a broad repertoire of antibodies, immunity to severe malaria targets a subset of conserved variants. These findings provide insights into antimalarial immunity and potential antibody biomarkers for disease risk.

Aim To determine the age specific immunity profile for rubella from three discrete study populations in Papua New Guinea, and to inform policy regarding the possible introduction of rubella vaccine. Background In 2005, the Western Pacific Region (WPR), of which Papua New Guinea (PNG) is a member state, declared the goal of regional measles elimination by 2012. Recently, WPR has incorporated an accelerated control goal for rubella and congenital rubella syndrome (CRS). PNG currently recommends two doses of measles vaccination at 6 and 9 months of age with a monovalent measles vaccine, which does not include rubella vaccine. Methods Convenience samples were collected from 1326 eligible participants in PNG and assessed for rubella immunity using the Dade Behring Enzygnost™ Anti-Rubella-Virus enzyme immunoassay. Nearly 34% were collected during an age stratified prospective survey of febrile patients in Madang Province; approximately 49% were collected from women of childbearing age in East Sepik and Milne Bay Provinces. Remaining specimens were collected from 6 to 7-month-old infants in Port Moresby prior to receiving the first dose of measles vaccine. Findings Of all samples tested, 65.2% (95% confidence interval (CI): 62.6–67.8) had evidence of immunity to rubella infection. Of women more than 15 years of age, 91.6% (95% CI: 89.4–93.5) were immune. The force of infection was highest between 5 and 19 years of age. Conclusions Although a population-based sample was not used, our multi-centre study of the population immunity profile suggests that immunity against rubella is extremely high in most women of childbearing age, but women who become pregnant at an early age may be at high risk of rubella infection during pregnancy and potential delivery of an infant with CRS. Routine measles vaccine coverage, a proxy for measles-rubella vaccine coverage, as measured in recently published studies, is well below the WHO target of 80% coverage. Introduction of a child or infant dose of rubella vaccine requires caution and further study.

Despite consistent evidence of a protective effect of alpha(+)-thalassemia against severe Plasmodium falciparum disease, the mechanisms underlying this protection remain unknown. An increase in risk of Plasmodium vivax malaria in early childhood resulting in a cross-species protection against severe P. falciparum malaria has been proposed as a possible mechanism in Melanesian children. The association of alpha(+)-thalassemia genotypes with a risk of P. falciparum and P. vivax infection and uncomplicated illness was reassessed in a cohort of 1,112 Papua New Guinean children, followed from 3-21 months of age. Three hundred and eighty-nine (35.0%) children were homozygous for a(+)-thalassemia (-a/-a), 506 (45.5%) heterozygous (aa/-a) and 217 (19.5%) homozygous for the wild-type allele. No significant differences in the incidence of P. falciparum (Pf) or P. vivax (Pv) malaria were observed between alpha(+)-thalassemia homozygote (Pf: Incidence rate ratio (IRR) = 1.13, CI(95) (0.82,1.56), P = 0.45, Pv: IRR = 1.15, CI(95) (0.9881.56), P = 0.31), heterozygote (Pf: IRR = 0.98, CI(95) (0.71,1.34), P = 0.93, Pv: IRR = 1.14, CI(95) (0.88,1.48), P =0.33) and wild-type children. The prevalence of infection with either species did not differ between alpha(+)-thalassemia genotypes, although densities of P. vivax (but not of P. falciparum) infections were significantly higher in alpha(+)-thalassemia homozygote and heterozygote children. An excessive risk of moderate-to-severe anemia (hemoglobin (Hb) > 8 g/dl) was observed in alpha(+)-thalassemia homozygote children (IRR = 1.54, CI(95)(1.12, 2.11), P = 0.008). This study therefore failed to confirm an increased risk of P. vivax or P. falciparum malaria in very young, alpha(+)-thalassemic children without significant levels of acquired immunity. This confirms the lack of protection by alpha(+)-thalassemia against uncomplicated P. falciparum and challenges the hypothesis of immunological cross-protection between P. falciparum and P. vivax as a mechanism underlying alpha(+)-thalassemia protection against severe P. falciparum disease in Melanesian children

Background Acute lower respiratory tract infections (ALRI) commonly result in fatal outcomes in the young children of Papua New Guinea (PNG). However, comprehensive studies of the viral aetiology of ALRI have not been conducted in PNG for almost 30 years. Objectives To determine the viruses associated with ALRI among children living in the PNG highlands using sensitive molecular detection techniques. Study design Pernasal swabs were collected routinely between 1 week and 18 months of age and also during episodes of ALRI, as part of a neonatal pneumococcal conjugate vaccine trial. A tandem multiplex real-time PCR assay was used to test for a comprehensive range of respiratory viruses in samples collected from 221 young children. Picornavirus typing was supported by DNA sequence analysis. Results Recognized pathogenic respiratory viruses were detected in 198/273 (73%) samples collected from children with no evidence of ALRI and 69/80 (86%) samples collected during ALRI episodes. Human rhinoviruses (HRV) species A, B and C were detected in 152 (56%) samples from non-ALRI children and 50 (63%) samples collected during ALRI episodes. Partial structural region sequences for two new species C rhinoviruses were added to the GenBank database. ALRI was associated with detection of adenovirus species B (p

Concerns about the risk of inducing immune deviation-associated “neonatal tolerance” as described in mice have restricted the widespread adoption of neonatal vaccination. The aim of this study was to demonstrate the immunological feasibility of neonatal pneumococcal conjugate vaccination (PCV) which could potentially protect high-risk infants in resource poor countries against severe pneumococcal disease and mortality in the early critical period of life. Papua New Guinean infants were randomized to be vaccinated with the 7-valent PCV (7vPCV) at birth, 1 and 2 months (neonatal group, n=104) or at 1, 2 and 3 months of age (infant group, n=105), or to not receive 7vPCV at all (control group, n=109). Analysis of vaccine responses at 3 and 9 months of age demonstrated persistently higher type-1 (IFN-γ) and type-2 (IL-5 and IL-13) T-cell responses to the protein carrier CRM197 and IgG antibody titres to 7vPCV serotypes in children vaccinated with 7vPCV according to either schedule as compared to unvaccinated children. In a comprehensive immuno-phenotypic analysis at 9 months of age, no differences in the quantity or quality of vaccine-specific T cell memory responses were found between neonatal vaccinations versus children given their first PCV dose at one month. Hospitalization rates in the first month of life did not differ between children vaccinated with PCV at birth or not. These findings demonstrate that neonatal 7vPCV vaccination is safe and not associated with immunological tolerance. Neonatal immunisation schedules should therefore be considered in high-risk areas where this may result in improved vaccine coverage and the earliest possible protection against pneumococcal disease and death.

We evaluated loop-mediated isothermal amplification end-point detection methods for Salmonella, Shigella, and Vibrio cholerae. Detection sensitivities were comparable to real-time PCR methods. The colorimetric dyes hydroxynaphthol blue and SYBR Green I showed increased sensitivity when compared to visual and automated turbidity readings. End-point colorimetric dyes promise great utility in developing settings.

We determined the relationship between the serum concentrations of matrix metalloproteinase-9 (MMP-9) and tissue inhibitors of metalloproteinases 1 (TIMP-1) in 33 patients with subacute sclerosing panencephalitis (SSPE) to investigate the function of the blood-brain-barrier (BBB) in SSPE. Serum MMP-9 and TIMP-1 levels were measured by ELISA. Serum MMP-9 levels and MMP-9/TIMP-1 ratios of SSPE patients in Papua New Guinea (n = 24), and those in Japan (n = 9) were significantly higher than the each control (MMP-9, p = 0.0390, and p = 0.0023, respectively; MMP-9/TIMP-1, p = 0.0319, and p = 0.0009, respectively). Serum MMP-9 levels and MMP-9/TIMP-1 ratios of SSPE patients with Jabbour stage III (n = 13) were significantly higher than those with Jabbour stage II (n = 18) (p = 0.003, and p = 0.0412, respectively). There were no significant differences of serum TIMP-1 levels between the SSPE patients and controls. High serum MMP-9 and MMP-9/TIMP-1 levels will promote brain invasion through the BBB by immunocompetent cells in the blood. Our findings suggest that the balance of serum MMP-9 and TIMP-1 levels modulate the inflammatory cascade of SSPE.

Background Subacute sclerosing panencephalitis (SSPE) is a rare progressive inflammatory disease characterized by the persistent infection of the brain by the measles virus. However, the immunological pathophysiology of SSPE is still unclear. Methods We measured the concentrations of interferon-γ (IFN-γ), tumor necrosis factor-α (TNF-α), interleukin-2 (IL-2), IL-4, IL-6, IL-10, and soluble TNF receptor 1 (sTNFR1) in the serum and cerebrospinal fluid (CSF) of 23 patients with SSPE in Papua New Guinea (PNG), a country with a high incidence of SSPE, and Japanese controls by cytometric bead array or ELISA. Results The serum IL-6 and IL-10 levels of SSPE patients were significantly higher than those of controls ( p = 0.0075, and p = 0.0019, respectively). The serum IL-6 and IL-10 levels of SSPE patients with fever were significantly higher than those without fever ( p = 0.0107, and p = 0.0006, respectively). The CSF IL-6 levels of SSPE patients were significantly higher than those of controls ( p = 0.0218). The CSF IL-6 levels of SSPE patients with myoclonic jerks were significantly higher than those without myoclonic jerks ( p = 0.0189). There were no differences in serum IFN-γ, TNF-α, IL-2, IL-4, and sTNFR1, or CSF IFN-γ, TNF-α, IL-2, IL-4, IL-10, and sTNFR1 levels between the affected patients and controls. Conclusion Our present study suggests that serum IL-6 and IL-10 levels are related to fever, and the CSF IL-6 level, myoclonic jerks, in SSPE patients in PNG.

Background The protective effect of Mycobacterium bovis BCG vaccination against infection and atopy varies between populations. Objective To identify differences in neonatal responses to BCG between diverse populations and study longitudinal associations with memory T-cell responses. Methods Cord blood mononuclear cells were collected from Papua New Guinean (PNG) and Western Australian (WA) newborns. Toll-like receptor (TLR)-2, TLR4, and TLR9 mRNA expression and in vitro BCG-stimulated (±IFN-γ priming) innate cytokine responses were compared. When PNG infants were 3 months old, PBMCs were stimulated in vitro with Mycobacterium -purified protein derivative (PPD) to determine memory T-cell responses. Results BCG-induced IL-10 and IFN-γ responses were significantly higher in cord blood mononuclear cells of PNG newborns, and TLR2 and TLR9 expression was significantly higher and TLR4 expression lower compared with WA newborns. High neonatal IL-10 and low IFN-γ responses to BCG were found to promote the development of PPD-memory T H 2 responses in infancy, whereas neonatal BCG-TNFα responses inhibited the development of PPD-IL 10 responses. When primed with IFN-γ, BCG-induced TNF-α, IL-12p70, and in particular IFN-γ responses were enhanced to a significantly higher extent in WA than in PNG newborns. In response to IFN-γ priming and BCG stimulation, natural killer cells of WA newborns produced IFN-γ, whereas natural killer cells of PNG newborns contributed only indirectly to this response. Conclusion Neonatal BCG-related innate immune responses control the differentiation of T H memory responses and vary between populations. This may explain differences in the effects of BCG vaccination between populations.