Helminth-host-environment interactions: Looking down from the tip of the iceberg.

In 1978, the theory behind helminth parasites having the potential to regulate the abundance of their host populations was formalized based on the understanding that those helminth macroparasites that reduce survival or fecundity of the infected host population would be among the forces limiting unregulated host population growth. Now, 45 years later, a phenomenal breadth of factors that directly or indirectly affect the host-helminth interaction has emerged. Based largely on publications from the past 5 years, this review explores the host-helminth interaction from three lenses: the perspective of the helminth, the host, and the environment. What biotic and abiotic as well as social and intrinsic host factors affect helminths? What are the negative, and positive, implications for host populations and communities? What are the larger-scale implications of the host-helminth dynamic on the environment, and what evidence do we have that human-induced environmental change will modify this dynamic? The overwhelming message is that context is everything. Our understanding of second-, third-, and fourth-level interactions is extremely limited, and we are far from drawing generalizations about the myriad of microbe-helminth-host interactions.Yet the intricate, co-evolved balance and complexity of these interactions may provide a level of resilience in the face of global environmental change. Hopefully, this albeit limited compilation of recent research will spark new interdisciplinary studies, and application of the One Health approach to all helminth systems will generate new and testable conceptual frameworks that encompass our understanding of the host-helminth-environment triad.

Parting ways: parasite release in nature leads to sex-specific evolution of defence.

We evaluated the extent to which males and females evolve along similar or different trajectories in response to the same environmental shift. Specifically, we used replicate experimental introductions in nature to consider how release from a key parasite (Gyrodactylus) generates similar or different defence evolution in male vs. female guppies (Poecilia reticulata). After 4-8 generations of evolution, guppies were collected from the ancestral (parasite still present) and derived (parasite now absent) populations and bred for two generations in the laboratory to control for nongenetic effects. These F2 guppies were then individually infected with Gyrodactylus, and infection dynamics were monitored on each fish. We found that parasite release in nature led to sex-specific evolutionary responses: males did not show much evolution of resistance, whereas females showed the evolution of increased resistance. Given that male guppies in the ancestral population had greater resistance to Gyrodactylus than did females, evolution in the derived populations led to reduction of sexual dimorphism in resistance. We argue that previous selection for high resistance in males constrained (relative to females) further evolution of the trait. We advocate more experiments considering sex-specific evolutionary responses to environmental change.

Impact of host sex and group composition on parasite dynamics in experimental populations.

To better understand the spread of disease in nature, it is fundamentally important to have broadly applicable model systems with readily available species which can be replicated and controlled in the laboratory. Here we used an experimental model system of fish hosts and monogenean parasites to determine whether host sex, group size and group composition (single-sex or mixed-sex) influenced host-parasite dynamics at an individual and group level. Parasite populations reached higher densities and persisted longer in groups of fish compared with isolated hosts and reached higher densities on isolated females than on isolated males. However, individual fish within groups had similar burdens to isolated males regardless of sex, indicating that females may benefit more than males by being in a group. Relative condition was positively associated with high parasite loads for isolated males, but not for isolated females or grouped fish. No difference in parasite dynamics between mixed-sex groups and single-sex groups was detected. Overall, these findings suggest that while host sex influences dynamics on isolated fish, individual fish in groups have similar parasite burdens, regardless of sex. We believe our experimental results contribute to a mechanistic understanding of host-parasite dynamics, although we are cautious about directly extrapolating these results to other systems.

Typhoid fever acquired in the United States, 1999-2010: epidemiology, microbiology, and use of a space-time scan statistic for outbreak detection.

Although rare, typhoid fever cases acquired in the United States continue to be reported. Detection and investigation of outbreaks in these domestically acquired cases offer opportunities to identify chronic carriers. We searched surveillance and laboratory databases for domestically acquired typhoid fever cases, used a space-time scan statistic to identify clusters, and classified clusters as outbreaks or non-outbreaks. From 1999 to 2010, domestically acquired cases accounted for 18% of 3373 reported typhoid fever cases; their isolates were less often multidrug-resistant (2% vs. 15%) compared to isolates from travel-associated cases. We identified 28 outbreaks and two possible outbreaks within 45 space-time clusters of ⩾2 domestically acquired cases, including three outbreaks involving ⩾2 molecular subtypes. The approach detected seven of the ten outbreaks published in the literature or reported to CDC. Although this approach did not definitively identify any previously unrecognized outbreaks, it showed the potential to detect outbreaks of typhoid fever that may escape detection by routine analysis of surveillance data. Sixteen outbreaks had been linked to a carrier. Every case of typhoid fever acquired in a non-endemic country warrants thorough investigation. Space-time scan statistics, together with shoe-leather epidemiology and molecular subtyping, may improve outbreak detection.

Concurrent nematode infection and pregnancy induce physiological responses that impair linear growth in the murine foetus.

This study examined concurrent stresses of nematode infection and pregnancy using pregnant and non-pregnant CD1 mice infected 3 times with 0, 50 or 100 Heligmosomoides bakeri larvae. Physiological, energetic, immunological and skeletal responses were measured in maternal and foetal compartments. Resting metabolic rate (RMR) was elevated by pregnancy, but not by the trickle infection. Energy demands during pregnancy were met through increased food intake and fat utilization whereas mice lowered their body temperature during infection. Both infection and pregnancy increased visceral organ mass and both altered regional bone area and mineralization. During pregnancy, lumbar mineralization was lower but femur area and mineralization were higher. On the other hand, infection lowered maternal femur bone area and this was associated with higher IFN-gamma in maternal serum of heavily infected pregnant mice. Infection also reduced foetal crown-rump length which was associated with higher amniotic fluid IL-1 beta.

Temporal dynamics of epidermal responses of guppies Poecilia reticulata to a sublethal range of waterborne zinc concentrations.

This study assessed the histological changes in the epidermis of guppies Poecilia reticulata induced by waterborne zinc (Zn). Laboratory-reared P. reticulata fry were maintained individually in separate vessels containing artificial water (8 microg l(-1) Zn) to which 0, 15, 30, 60 or 120 microg l(-1) Zn was added. Their epidermal response to Zn was monitored regularly over 4 weeks. Compared with controls, mucus was rapidly released and mucous cell numbers decreased at all concentrations. Thereafter mucous release, epidermal thickness, numbers and size of mucous cells fluctuated at a rate that varied with Zn concentration, but fluctuations declined after day 18. Results clearly highlight the dynamic nature of the epidermal response to sublethal concentrations of waterborne Zn. In general, low concentrations of Zn induced a rapid response with reduced numbers and size of mucous cells and shift in mucin composition, and a subsequent thickening of the epidermis. Epidermal thickness and mucous cell area fluctuated over time but were normal after a month of exposure to low Zn concentrations. The number of mucous cells, however, remained low. Virtually all mucous cells from fish maintained in 15 and 60 microg l(-1) Zn contained acidic mucins throughout the month, whereas fish maintained at 30 microg l(-1) Zn responded by production of neutral mucins during the first 12 days followed by a mixture of neutral and acidic mucins. At 120 microg l(-1) Zn, the most dramatic effects were the gradual but sustained decrease in numbers and area of mucous cells, and the shift to acidic mucins in these cells. Thus, as concentration of Zn increased, the epidermal responses indicated a disturbed host response (dramatic decline in mucous cell numbers, with mixed composition of mucins), which may have been less effective in preventing Zn uptake across the epithelium.

Species composition and distribution of adult Anopheles (Diptera: Culicidae) in Panama.

Anopheles (Diptera: Culicidae) species composition and distribution were studied using human landing catch data over a 35-yr period in Panama. Mosquitoes were collected from 77 sites during 228 field trips carried out by members of the National Malaria Eradication Service. Fourteen Anopheles species were identified. The highest average human biting rates were recorded from Anopheles (Nyssorhynchus) albimanus (Wiedemann) (9.8 bites/person/night) and Anopheles (Anopheles) punctimacula (Dyar and Knab) (6.2 bites/person/night). These two species were also the most common, present in 99.1 and 74.9%, respectively, of the sites. Anopheles (Nyssorhynchus) aquasalis (Curry) was encountered mostly in the indigenous Kuna Yala Comarca along the eastern Atlantic coast, where malaria case history and average human biting rate (9.3 bites/person/night) suggest a local role in malaria transmission. An. albimanus, An. punctimacula, and Anopheles (Anopheles) vestitipennis (Dyar and Knab) were more abundant during the rainy season (May-December), whereas An. aquasalis was more abundant in the dry season (January-April). Other vector species collected in this study were Anopheles (Kerteszia) neivai (Howard, Dyar, and Knab) and Anopheles (Anopheles) pseudopunctipennis s.l. (Theobald). High diversity of Anopheles species and six confirmed malaria vectors in endemic areas of Panama emphasize the need for more detailed studies to better understand malaria transmission dynamics.

Inhibition of human factor VIIIa by anti-A2 subunit antibodies.

Human inhibitory alloantibodies and autoantibodies to Factor VIII (FVIII) are usually directed toward the A2 and/or C2 domains of the FVIII molecule. Anti-C2 antibodies block the binding of FVIII to phospholipid, but the mechanism of action of anti-A2 antibodies is not known. We investigated the properties of a patient autoantibody, RC, and a monoclonal antibody, 413, that bind to the region which contains the epitopes of all anti-A2 alloantibodies or autoantibodies studied to date. mAb 413 and RC were noncompetitive inhibitors of a model intrinsic Factor X activation complex (intrinsic FXase) consisting of Factor IXa, activated FVIII (FVIIIa), and synthetic phospholipid vesicles, since they decreased the Vmax of intrinsic FXase by > 95% at saturating concentrations without altering the Km. This indicates that RC and mAb 413 either block the binding of FVIIIa to FIXa or phospholipid or interfere with the catalytic function of fully assembled intrinsic FXase, but they do not inhibit the binding of the substrate Factor X. mAb 413 did not inhibit the increase in fluorescence anisotropy that results from the binding of Factor VIIIa to fluorescein-5-maleimidyl-D-phenylalanyl-prolyl-arginyl-FIXa (Fl-M-FPR-FIXa) on phospholipid vesicles in the absence of Factor X, indicating it does not inhibit assembly of intrinsic FXase. Addition of Factor X to Fl-M-FPR-FIXa, FVIIIa, and phospholipid vesicles produced a further increase in fluorescence anisotropy and a decrease in fluorescence intensity. This effect was blocked completely by mAb 413. We conclude that anti-A2 antibodies inhibit FVIIIa function by blocking the conversion of intrinsic FXase/FX complex to the transition state, rather than by interfering with formation of the ground state Michaelis complex.

Conditions under Which the Mean Fertility Is Maximized When a Population Is at a Stable Equilibrium.

It is assumed that there is a population with two alleles at one locus, random mating of adults and selection only involving differential fertilities. By making use of the Kuhn-Tucker theory of optimization under constraints, conditions are derived under which stable equilibrium frequencies x, y and z of the three genotypes are the same as those that maximize the mean fertility of the population. We derive all sets of frequencies of this type for the Hadeler-Liberman symmetric fertility model and all such sets for which at least one genotype is missing for the general model. If the population has frequencies that are initially near those at which there is both a stable equilibrium and maximization of the mean fertility, then the mean fertility phi(t) at time t is nondecreasing with t as t -> &. It is found that it is possible for the stable equilibrium maximum points (x, y, z) to be one or two points on a ridge on which the mean fertility phi is maximized or the entire set of points on the ridge. Furthermore, phi may be smaller on this ridge than at another stable equilibrium point at which phi is not even locally maximized.

Genetic variability of the beta-tubulin genes in benzimidazole-susceptible and -resistant strains of Haemonchus contortus.

Benzimidazole anthelmintics are the most common chemotherapeutic agents used to remove intestinal helminths from farm animals. The development of drug resistance within helminth populations is wide-spread and can render these drugs essentially useless. The mechanism of benzimidazole resistance appears to be common to many species ranging from fungi to nematodes and involves alterations in the genes encoding beta-tubulin. During the selection process resulting in resistance, there must be quantitative changes in the population gene pool. Knowledge of these changes would indicate the mechanisms underlying the spread of resistance in the population, which in turn could be used to design more effective drug administration strategies. To this end we have identified allelic variation at two beta-tubulin genes in Haemonchus contortus using restriction map analysis of individual adults. Extremely high levels of variation were identified at both loci within a susceptible strain. In two independently derived benzimidazole resistant strains, allele frequencies at both loci were significantly different from the susceptible strain but not from each other. The same alleles at both loci, in both resistant strains, were favored by selection with benzimidazoles, suggesting that both loci are involved in determining benzimidazole resistance. These data confirm that changes in allele frequency, rather than novel genetic rearrangements induced by exposure to the drug, explain the changes associated with benzimidazole resistance. These results also show that any DNA based test for the development of benzimidazole resistance must take into account the frequency of alleles present in the population and not simply test for the presence or absence of specific allelic types.

In vivo decrease in the expression of complement receptor 2 on B-cells in HIV infection.

OBJECTIVE: To investigate changes in the expression of complement receptor 2 (CR2) on B-cells from HIV-infected individuals. CR2 is the C3d/Epstein-Barr virus receptor and has been implicated in B-cell activation. Changes in its level of expression may therefore be associated with B-cell dysfunction. DESIGN: Cross-sectional study of HIV-infected adults and age-matched control donors. METHODS: The percentage expression and mean fluorescence intensity of CR2 (and three additional markers: CD19, CD69, and a standard antigen designation: HLA-DR) was measured on CD20+ B-cells using a two-color flow cytometric assay. RESULTS: This study demonstrated a highly significant (P = 0.0001) decrease in the percentage co-expression of CR2 on CD20+ B-cells in HIV-infected individuals, compared with control donors. The mean percentage of CD20+ cells co-expressing CR2 was 71% (s.d., +/- 15%) in the HIV-seropositive patients and 94% (s.d., +/- 4%) in the control group. The pattern of CR2 expression in a number of the patients suggested a decrease in antigen density on the cells. Decreased expression of CR2 did not correlate with disease stage (asymptomatic, AIDS-related complex, or AIDS), nor with CD4+ T-cell percentage or absolute count, in the seropositive group. CONCLUSIONS: The evidence for a role for CR2 in B-cell activation suggests that its decreased expression, which we have demonstrated in HIV-seropositive individuals, may be associated with the B-cell dysfunction observed in HIV infection. Our finding that expression of this marker is decreased even in asymptomatic patients is consistent with reports of early B-cell defects in such individuals. Further investigation of this possible association may shed some light on both the increased incidence of bacterial infections in HIV-infected adults and children and their impaired responses to certain immunizations.

Single-column purification of free recombinant proteins using a self-cleavable affinity tag derived from a protein splicing element.

A novel protein purification system has been developed which enables purification of free recombinant proteins in a single chromatographic step. The system utilizes a modified protein splicing element (intein) from Saccharomyces cerevisiae (Sce VMA intein) in conjunction with a chitin-binding domain (CBD) from Bacillus circulans as an affinity tag. The concept is based on the observation that the modified Sce VMA intein can be induced to undergo a self-cleavage reaction at its N-terminal peptide linkage by 1,4-dithiothreitol (DTT), beta-mercaptoethanol (beta-ME) or cysteine at low temperatures and over a broad pH range. A target protein is cloned in-frame with the N-terminus of the intein-CBD fusion, and the stable fusion protein is purified by adsorption onto a chitin column. The immobilized fusion protein is then induced to undergo self-cleavage under mild conditions, resulting in the release of the target protein while the intein-CBD fusion remains bound to the column. No exogenous proteolytic cleavage is needed. Furthermore, using this procedure, the purified free target protein can be specifically labeled at its C-terminus.

Inhibitors of human purine nucleoside phosphorylase. Synthesis, purine nucleoside phosphorylase inhibition, and T-cell cytotoxicity of 2,5-diaminothiazolo[5,4-d]pyrimidin-7(6H)-one and 2,5-diaminothiazolo[4,5-d]pyrimidin-7(6H)-one. Two thio isosteres of 8-aminoguanine.

8-Aminoguanine is a potent inhibitor of purine nucleoside phosphorylase (PNP) and also a substrate of PNP. Two thio isosteres of 8-aminoguanine, 2,5-diaminothiazolo[5,4-d]pyrimidin-7(6H)-one (2) and 2,4-diaminothiazolo[4,5-d]pyrimidin-7(6H)-one (3), which cannot be substrates of PNP, were synthesized and evaluated for their inhibitory activity against PNP. They were found to be weak inhibitors of PNP and to be noncytotoxic for MOLT-4 T-cells in culture.

Physical studies on isolated human prothrombin fragment-2. Comparisons with human prothrombin fragment-1.

Variation of pH strongly affects the fluorescence intensity of human prothrombin fragment-1 in a manner suggesting contributions from a number of protropic equilibria including groups with apparent pKa values near 3.0. These results suggest a structural role for pK1a of gamma-carboxyglutamic acid noieties. Added calcium ions (9 mM calcium chloride) quench the fluorescence titration curve uniformly above pH 4. Below pH 4, however, the titration curve in the presence of calcium ions suggests that calcium-ion-dependent processes leading to fluorescence quenching are pH-dependent. Upon back titration of human fragment-1, from pH 9, hysteresis is observed. Human prothrombin fragment-2 fluorescence titration curves are relatively broad at low pH suggesting the titration of normal carboxyl groups. The titration curves of fragment-2 are not affected by the presence of calcium ions, and hysteresis occurs upon back titration from low pH values. Circular dichroism (CD) Cotton effects appear at 232 nm and 280 nm and a trough appears at 203 nm in the CD spectrum of human prothrombin fragment-2. The Cotton effects in the region from 230 nm to 300 nm are sensitive to pH, ellipticity values at 232 nm increasing from approximately 300 at pH 2.5 to 1300 (degree-cm/decimole) at neutral pH and finally become negative at high pH values. In contrast to fragment-1, at neutral pH the fragment-2 Cotton effect at 232 nm is insensitive to the presence of 8 mM calcium chloride.

Relation of coronary arterial patency and left ventricular function to electrocardiographic changes after streptokinase treatment during acute myocardial infarction.

One hundred and eighty-eight patients with evolving acute myocardial infarction were treated with intravenous streptokinase. Serial 12-lead electrocardiograms were recorded for 3 hours after treatment and inspected for rapid repolarization changes of the ST segment and T wave. Abrupt electrocardiographic repolarization changes were observed in 106 patients (56%) and were strongly predictive for an open infarct-related coronary artery at a mean of 6 days after treatment (predictive value = 0.92, sensitivity = 0.67). Abrupt electrocardiographic changes were not observed in 82 patients (44%). This absence was not a good predictor of an occluded infarct-related coronary artery (predictive value = 0.4). There was no relation between the presence or absence of abrupt electrocardiographic changes and global or regional left ventricular function after streptokinase treatment. Abrupt repolarization changes after thrombolytic treatment indicate a high probability of an open infarct-related artery. When abrupt repolarization changes do not occur, the patency of the infarct-related coronary artery cannot be predicted with accuracy. Serial electrocardiographic recordings do not provide sufficient information about coronary patency to eliminate the need for coronary arteriography.

Life-history variation among lines isolated from a laboratory population of Heligmosomoides polygyrus bakeri.

The extent of variation in several life-history traits within a laboratory population of the parasitic nematode, Heligmosomoides polygyrus bakeri, was studied in 10 relatively inbred parasite lines isolated from a stock population and characterized in BALB/c mice after 4, 8 and 11 generations of isolation. As expected, within-line variation for most traits at generation 11 compared to generation 4 significantly decreased (P < 0.01). At each generation of characterization, variation was observed among lines for parasite establishment, rate of development in the host, rate of early egg production, per capita fecundity, short-term and long-term survival and profiles of egg production, rate of decline in egg production, life-long reproductive effort and in vitro egg hatchability. Measures of all traits, except establishment, were highly repeatable. The rate of development was higher at generation 8 compared to the stock (P < 0.0001), and regression analysis revealed that early egg production of lines increased over 11 generations of isolation (P = 0.003). These results, together with the observed decrease (P < 0.01) in total variation of most of the traits over all lines during the process of isolation, suggested an evolutionary response of the traits, probably to the rapid passage of lines every month. The rate of development subsequently decreased between generations 8 and 11 in all lines (P < 0.0001), suggesting that the random genetic drift procedure used to isolate the lines eventually exerted detrimental inbreeding effects on this trait. The evolutionary responses of life-history traits to rapid passage and inbreeding suggest a genetic basis for variation in these life-history traits.

The efficacy of ivermectin against laboratory strains of Heligmosomoides polygyrus (Nematoda).

A susceptible strain of Heligmosomoides polygyrus was selected for 15 generations with increasing doses of ivermectin (0-6 mg/kg). A passage strain was developed, parallel with the ivermectin-selected strain, to control for changes due to rapid passage from mouse to mouse. The LD50s of the 8th and 15th generations of the ivermectin-selected strain were 1.5 times that of the susceptible strain. The LD50 of the passage strain at generations 8 and 15 remained similar to that of the susceptible strain. Ivermectin efficacy was lower against the LA stage than against the adult stage in the susceptible strain, the Ivermectin-selected strain and the passage strain at generation 8.

Age-related differences in prevalence and intensity of Trichuris trichiura in sensory impaired children from two schools in Guatemala.

A survey of 102 blind and 218 deaf children in 2 specialized schools in Guatemala revealed distinctly different age-intensity profiles of infection with the intestinal helminth, Trichuris trichiura. In the school for the deaf, intensity (measured as eggs per gram of stool) peaked in the children 7 to 8 years old and dropped to very low average levels in the teenagers. By contrast, in the school for the blind, the intensity of T. trichiura remained low until the early teens. Intensity peaked in the age group 13 to 14 years old, and then decreased in the 15 to 17 years old pupils. Behavioural differences between blind and deaf children, or differences between the schools, may have contributed to this differential pattern of infection.

Clustering of Ascaris lumbricoides and Trichuris trichiura infections within households.

A survey of 428 households in a shanty town in Coatzacoalcos, Mexico, revealed high prevalences of Ascaris lumbricoides and Trichuris trichiura. The data were analysed separately for A. lumbricoides and T. trichiura in order to investigate the spatial distribution of heavy infections through the town. Within each age class, those individuals with egg counts in the upper 20% of the range for that age were classified as "heavily infected". When the data were stratified by household size, it was found that the distribution of "heavily-infected" individuals was not random in the community. "Heavily-infected" individuals were found together in households; fewer household units had a single heavily-infected individual than would be expected by chance. Such a pattern could result either from genetic similarities among family members influencing their ability to mount an effective immunological response to infection, or focal transmission in the vicinity of the home, or both. This result may have important implications for the development of community control programmes.

Predisposition of individuals and families in Mexico to heavy infection with Ascaris lumbricoides and Trichuris trichiura.

Stool specimens from 90% of the population of 2 marginal communities in Coatzacoalcos, Mexico, were examined by the Kato-Katz technique for the estimation of eggs per gram of faeces (epg). Average epg values for Ascaris lumbricoides and Trichuris trichiura were calculated for each family in the community, based on the arithmetic mean epg of family members for each parasite. 39 families were classified as either lightly or heavily infected with A. lumbricoides and infected members were treated with mebendazole. A. lumbricoides expelled following treatment were collected from the children 2-10 years of age, and reinfection was monitored monthly for 6 months. At the end of the reinfection interval, a final stool specimen was collected from all family members and those infected were treated. Again, A. lumbricoides expelled by the children 2-10 years of age were collected. There was a significant correlation between the number of A. lumbricoides expelled by the children at the first and second treatments as well as in epg values before the first and second treatments for both A. lumbricoides and T. trichiura in children (2-10 years) and adults (greater than 19 years). At the family level, there were significant correlations between the values of family mean epg before each treatment for both A. lumbricoides and T. trichiura.