The endoplasmic reticulum (ER) plays a critical role in the synthesis and chaperoning of membrane-associated and secreted proteins. The membrane is also an important site of Ca(2+) storage and release. Calreticulin is a unique ER luminal resident protein. The protein affects many cellular functions, both in the ER lumen and outside of the ER environment. In the ER lumen, calreticulin performs two major functions: chaperoning and regulation of Ca(2+) homoeostasis. Calreticulin is a highly versatile lectin-like chaperone, and it participates during the synthesis of a variety of molecules, including ion channels, surface receptors, integrins and transporters. The protein also affects intracellular Ca(2+) homoeostasis by modulation of ER Ca(2+) storage and transport. Studies on the cell biology of calreticulin revealed that the ER membrane is a very dynamic intracellular compartment affecting many aspects of cell physiology.

Calreticulin: one protein, one gene, many functions.

The threat posed by the highly pathogenic H5N1 influenza virus requires public health authorities to prepare for a human pandemic. Although pre-pandemic vaccines and antiviral drugs might significantly reduce illness rates, their stockpiling is too expensive to be practical for many countries. Consequently, alternative control strategies, based on non-pharmaceutical interventions, are a potentially attractive policy option. School closure is the measure most often considered. The high social and economic costs of closing schools for months make it an expensive and therefore controversial policy, and the current absence of quantitative data on the role of schools during influenza epidemics means there is little consensus on the probable effectiveness of school closure in reducing the impact of a pandemic. Here, from the joint analysis of surveillance data and holiday timing in France, we quantify the role of schools in influenza epidemics and predict the effect of school closure during a pandemic. We show that holidays lead to a 20-29% reduction in the rate at which influenza is transmitted to children, but that they have no detectable effect on the contact patterns of adults. Holidays prevent 16-18% of seasonal influenza cases (18-21% in children). By extrapolation, we find that prolonged school closure during a pandemic might reduce the cumulative number of cases by 13-17% (18-23% in children) and peak attack rates by up to 39-45% (47-52% in children). The impact of school closure would be reduced if it proved difficult to maintain low contact rates among children for a prolonged period.

Estimating the impact of school closure on influenza transmission from Sentinel data

Malaria remains a major cause of morbidity and mortality in the tropics, with Plasmodium falciparum responsible for the majority of the disease burden and P. vivax being the geographically most widely distributed cause of malaria. Gametocytes are the sexual-stage parasites that infect Anopheles mosquitoes and mediate the onward transmission of the disease. Gametocytes are poorly studied despite this crucial role, but with a recent resurgence of interest in malaria elimination, the study of gametocytes is in vogue. This review highlights the current state of knowledge with regard to the development and longevity of P. falciparum and P. vivax gametocytes in the human host and the factors influencing their distribution within endemic populations. The evidence for immune responses, antimalarial drugs, and drug resistance influencing infectiousness to mosquitoes is reviewed. We discuss how the application of molecular techniques has led to the identification of submicroscopic gametocyte carriage and to a reassessment of the human infectious reservoir. These components are drawn together to show how control measures that aim to reduce malaria transmission, such as mass drug administration and a transmission-blocking vaccine, might better be deployed.

Epidemiology and Infectivity of Plasmodium falciparum and Plasmodium vivax Gametocytes in Relation to Malaria Control and Elimination

Parasitic nematodes that cause elephantiasis and river blindness threaten hundreds of millions of people in the developing world. We have sequenced the ∼90 megabase (Mb) genome of the human filarial parasite Brugia malayi and predict ∼11,500 protein coding genes in 71 Mb of robustly assembled sequence. Comparative analysis with the free-living, model nematode Caenorhabditis elegans revealed that, despite these genes having maintained little conservation of local synteny during ∼350 million years of evolution, they largely remain in linkage on chromosomal units. More than 100 conserved operons were identified. Analysis of the predicted proteome provides evidence for adaptations of B. malayi to niches in its human and vector hosts and insights into the molecular basis of a mutualistic relationship with its Wolbachia endosymbiont. These findings offer a foundation for rational drug design.

Draft genome of the filarial nematode parasite Brugia malayi (Science (2007) (1756))

Plasmodium vivax is a major cause of febrile illness in endemic areas of Asia, Central and South America, and the horn of Africa. Plasmodium vivax infections are characterized by relapses of malaria arising from persistent liver stages of the parasite (hypnozoites) which can be prevented only by 8-aminoquinoline anti-malarials. Tropical P. vivax relapses at three week intervals if rapidly eliminated anti-malarials are given for treatment, whereas in temperate regions and parts of the sub-tropics P. vivax infections are characterized either by a long incubation or a long-latency period between illness and relapse - in both cases approximating 8-10 months. The epidemiology of the different relapse phenotypes has not been defined adequately despite obvious relevance to malaria control and elimination. The number of sporozoites inoculated by the anopheline mosquito is an important determinant of both the timing and the number of relapses. The intervals between relapses display a remarkable periodicity which has not been explained. Evidence is presented that the proportion of patients who have successive relapses is relatively constant and that the factor which activates hypnozoites and leads to regular interval relapse in vivax malaria is the systemic febrile illness itself. It is proposed that in endemic areas a large proportion of the population harbours latent hypnozoites which can be activated by a systemic illness such as vivax or falciparum malaria. This explains the high rates of vivax following falciparum malaria, the high proportion of heterologous genotypes in relapses, the higher rates of relapse in people living in endemic areas compared with artificial infection studies, and, by facilitating recombination between different genotypes, contributes to P. vivax genetic diversity particularly in low transmission settings. Long-latency P. vivax phenotypes may be more widespread and more prevalent than currently thought. These observations have important implications for the assessment of radical treatment efficacy and for malaria control and elimination.

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Determinants of relapse periodicity in Plasmodium vivax malaria.

ABSTRACr: The population dynamics of cultured Plasmodium falciparum parasites was examined during their sporogonic development in Anopheles gambiae mosquitoes. Estimates of absolute denisities were determined for each life stage, and life tables were contructed for each of 38 experimental infections. Macrogametocyte and ookinete mortalities contributed equally to the overall mortality. On average, there was a 40-fold decrease in parasite numbers in the transition from the macrogametocyte to the ookinete stage, a 69-fold decrease in the transition from ookinete to oocyst stages, and a total net decrease in parasite numbers from macrogametocyte to oocyst stage of 2,754-fold (i.e., multiplicative). There was no relationship between macrogametocyte and ookinete densities due to the inherent variability in fertility among different gametocyte cultures. There was a curvilinear relationship (r2 = 0.66) between ookinete and oocyst densities. Above a threshold of about 30 ookinetes/mosquito, the oocyst yield per ookinete became increasingly greater with increasing ookinete density. There was a linear relationship (r2 = 0.73) between oocyst and sporozoite densities, with an average of 663 salivary gland sporozoites produced per oocyst. Sporozoite production per oocyst was not affected by oocyst density and virtually all oocyst infections resulted in sporozoite infections of the salivery glands. This quantitative study indicates that the sporogony of cultured P. falciparum in laboratory-infected A. gambiae is an inefficient process and that the ookinete is the key transitional stage affecting the probability of vector infectivity.

Sporogony in laboratory-infected anopheles gambiae

Abstract Systemic parasiticides in livestock can control zoophilic malaria vectors that contribute to residual malaria transmission. Membrane feeding techniques were used to screen seven systemic parasiticidic drugs currently in veterinary use for livestock and dogs. Drugs were tested in two laboratory strains of zoophilic Anopheles –A. stephensi (South Asian vector) and A. albimanus (Central American vector). To assess the relative potentials of these drugs, the resultant LC-50 for each drug was compared with what is known about the pharmacokinetic of the drug. Drugs with LC-50 values below the reported maximum plasma concentration of treated animals were considered as showing the most promise for use in the field. Ivermectin and fipronil showed the greatest promise for use in cattle against A. stephensi. Fipronil showed the greatest promise for use in cattle against A. albimanus. Both fluralaner and afoxolaner were highly effective against both mosquito species but pharmacokinetic data for these drugs in cattle are lacking. Eprinomectin, moxidectin and abamectin showed marginal to no promise for either mosquito species. At sublethal doses, ivermectin, fipronil, and afoxolaner (but not fluralaner) significantly reduced the larval production of surviving A. stephensi and A. albimanus. Further testing of candidate systemic parasiticides, including their product formulations, in livestock against field-collected populations of Anopheles is the next logical step toward full implementation of this strategy to manage zoophilic vectors.

Survival and Fecundity of Anopheles stephensi and Anopheles albimanus Mosquitoes (Diptera: Culicidae) After Ingesting Bovine Blood Containing Various Veterinary Systemic Parasiticides

The STECLA strain of Anopheles albimanus has been in continuous colony for many years and is the reference strain on which genomic studies for the species are based. Recently, the STECLA strain was demonstrated to be much less susceptible to ivermectin ingested in a blood meal (4-day LC50 of 1468 ng/ml) than all other Anopheles species tested to-date (LC50 values range from 7 to 56 ng/ml). The ability of An. albimanus to survive ingestion of ivermectin at concentrations far beyond that typically found in the blood of ivermectin-treated people or livestock (i.e., 30-70 ng/ml) could invalidate the use of ivermectin as a malaria vector control strategy in areas where An. albimanus is a primary vector.To investigate this, host-seeking An. albimanus were captured in northern Belize and used in membrane feeding bioassays of ivermectin, employing the same methods as described earlier with the STECLA strain.Field-collected An. albimanus in Belize were 55 times more susceptible to ingested ivermectin than were the STECLA reference strain. Oral susceptibility to ivermectin in wild An. albimanus from Belize (4-day LC50 of 26 ng/ml) was equivalent to that of other Anopheles species tested.Contrary to initial assessments using a highly inbred strain of mosquito, laboratory studies using a field population indicate that ivermectin treatment of livestock could reduce An. albimanus populations in areas of Central America and the Caribbean where malaria transmission may occur. Toxicity screening of ivermectin and other systemic parasiticides for malaria control should examine wild populations of the vector species being targeted. 

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Oral susceptibility to ivermectin is over fifty times greater in a wild population of Anopheles albimanus mosquitoes from Belize than the STECLA laboratory reference strain of this mosquito

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The transmission cycle of West Nile virus (WNV) involves multiple species of birds. The relative importance of various bird species to the overall transmission is often inferred from the level and duration of viremia that they experience upon infection. Reports utilizing in vitro feeding techniques suggest that the source and condition of blood in which arboviruses are fed to mosquitoes can significantly alter the infectiousness of arbovirus to mosquitoes. We confirmed this using live hosts. A series of mosquito feedings with Culex pipiens was conducted on WNV-infected American robins and common grackles over a range of viremias. Mosquitoes were assayed individually by plaque assay for WNV at 3 to 7 days after feeding. At equivalent viremia, robins always infected more mosquitoes than did grackles. We conclude that the infectiousness of viremic birds cannot always be deduced from viremia alone. If information concerning the infectiousness of a particular bird species is important, such information is best acquired by feeding mosquitoes directly on experimentally infected individuals of that species.

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Jefferson A. Vaughan

Papers

Sporogony in laboratory-infected anopheles gambiae

Survival and Fecundity of Anopheles stephensi and Anopheles albimanus Mosquitoes (Diptera: Culicidae) After Ingesting Bovine Blood Containing Various Veterinary Systemic Parasiticides

Oral susceptibility to ivermectin is over fifty times greater in a wild population of Anopheles albimanus mosquitoes from Belize than the STECLA laboratory reference strain of this mosquito

Oral susceptibility to ivermectin is over fifty times greater in a wild population of Anopheles albimanus mosquitoes from Belize than the STECLA laboratory reference strain of this mosquito

Bird species define the relationship between West Nile viremia and infectiousness to Culex pipiens mosquitoes