The effects of gamma-irradiation on migration and survival of Schistosoma mansoni schistosomula in mice

Infections with irradiated Schistosoma mansoni were established by intramuscular (i.m.) injection of mechanically transformed schistosomula. A dose of 2.3 krad. allowed persistence of a small proportion of worms to adulthood, and of these survivors the majority of the female worms were sexually sterile. However, a small proportion of 2.3 krad.-irradiated females and a larger proportion of similarly irradiated males were capable of pairing successfully with non-irradiated partners. Radiation in the range 2.3 to 10 krad. resulted in slightly reduced peak recoveries from the lungs while 20 krad. resulted in a much reduced and 40 krad. a virtual absence of survival to the lung stage. Increasing doses of radiation in the range 2.3 to 10 krad. resulted in successively fewer parasites reaching the liver. Thus, the major sites of the radiation-induced mortality appeared to be as follows: 2.3 krad., mainly in the liver; 4 krad., in the lungs and liver; 10 krad., mainly in the lungs; 20 krad., at the injection site and in the lungs and 40 krad., mainly at the injection site. The infections studied here showed reduced survival following exposure to high doses of radiation compared with the infections, established as percutaneously applied cercariae, which have been reported by other workers. Possible reasons for the disparity are discussed.     

Schistosoma mansoni: Interactive effects of irradiation and cryopreservation on parasite maturation and immunization of mice

Mechanically transformed schistosomula of Schistosoma mansoni were irradiated with levels of ⁶⁰Co irradiation between 2.5 and 54 krad, cryopreserved by the two-step addition of ethanediol and rapid cooling technique, and were injected intramuscularly into groups of mice which were perfused 40 days later. The schistosomula were either irradiated and then cryopreserved (IC) or cryopreserved and then irradiated in the frozen state (CI). Development into adult worms was prevented with 4 krad for IC schistosomula, but for CI schistosomula a small number of worms (1.6%) was recovered using 8.8 krad. A dose of 4 krad was sufficient to prevent development of unfrozen controls (I), but for schistosomula irradiated while exposed to ethanediol (EI), a dose of 7 krad was required. Using the different protocols, the peak levels of protection against a challenge infection were achieved with 9 (IC) and 16 krad (CI), compared to 20 krad for unfrozen schistosomula (I) reported previously. The highest level of protection (65%) was achieved with CI schistosomula. Possible interactions between the radioprotective and damaging effects of cryopreservation are discussed.     

Immunization of zebu calves against Fasciola gigantica, using irradiated metacercariae

The pathogenesis of unirradiated, 3 krad-irradiated and 20 krad-irradiated metacercarial infections was compared in zebu calves studied over a 10-week period. Calves exposed to 1000 unirradiated metacercariae (mc) became hypoalbuminaemic, and showed elevated serum concentrations of liver enzymes, whereas neither of the other groups was significantly affected. At slaughter, a mean of 332 flukes was recovered from the 0 krad group, while only 23% and 12% of this number were recovered from the 3 krad and and 20 krad groups, respectively. All the worms recovered from the 20 krad group were stunted, and found in biliary ductules, but a mean of 13% of the flukes recovered from the 3 krad group were large, and dwelling in main bile-ducts. Liver lesions typical of acute fascioliasis were present in the 0 krad group, but lesions in the other groups, and particularly the 20 krad group, were far less severe. Judged on clinico-pathological criteria, a single vaccination of calves with 1000 3 krad-irradiated mc induced partial resistance to a challenge with 1000 normal mc eight weeks later, but the reduction in worm recovery was not statistically significant. There was less evidence of protection when two vaccinating doses of 3 krad mc were given within four weeks, with challenge at week 8, and a single vaccination was ineffective against a challenge four weeks later. However, when the irradiation dose was increased to 20 krad, a hgh level of resistance (69% worm reduction) was induced by a single vaccination, given eight weeks before challenge, and liver pathology was strikingly reduced in the vaccinated animals.     

Schistosoma mansoni: Infectivity and immunizing effects of in vitro derived schistosomula attenuated by X irradiation

Irradiated and nonirradiated in vitro derived schistosomula of Schistosoma mansoni were injected intraperitoneally into mice. Sixteen percent of nonirradiated schistosomula, 8% of those irradiated with 1000 R, and virtually none of those irradiated with 3000 R and above survived in mice for 5 weeks. However, those irradiated with 3000 R survived in small numbers for shorter periods of time. Schistosomula irradiated with 3000 or 6000 R were used to immunize mice against subsequent infection with cercariae. Prior ip injections of schistosomula irradiated with 3000 R resulted in reductions in worm burden after challenge from 5 to 91%; the observed protection was related to the number of inoculations. The subcutaneous route appeared to be less effective. Schistosomula irradiated with 6000 R produced less protection than those irradiated with 3000 R.     

Schistosoma mansoni: immunization of cynomolgus monkeys by injection of irradiated schistosomula.

Although the immunization of primates with irradiated schistosome cercariae has been demonstrated, no success has been reported by injection with the irradiated schistosomule stage. The present investigation was designed to test whether cynomolgus monkeys could be protectively immunized with ⁶⁰Co-irradiated Schistosoma mansoni schistosomula. Monkeys injected once with 10⁴ irradiated schistosomula (50 krad at 4 krad/min) had 52% fewer challenge worms than the control group at necropsy. Four immunizations did not induce a higher level of resistance. At 50 days post-challenge, the immunized monkeys excreted 80% fewer eggs than did the control animals. An attempt to enhance irradiated schistosomule-induced protection with tetramisole · HCl was unsuccessful.     

Immunisation of sheep against Schistosoma mattheei using either irradiated cercariae or irradiated schistosomula.

Irradiated cercariae, irradiated schistosomula, or heterologous infections were used to vaccinate sheep against Schistosoma mattheei infection. In the first experiment four doses of 10(4) S. mattheei cercariae irradiated at 6Kr were administered to sheep by percutaneous infection at 4 week intervals. This induced a 74% reduction in a challenge infection compared to control sheep while only 13% protection was achieved in a third group of sheep immunised with normal cercariae of the heterologous parasite S. mansoni. No significant differences were seen in histopathology of the liver of any of the sheep but the pathological changes were more severe in the large and small intestines of sheep vaccinated with the heterologous parasite. In the second experiment with irradiated cercariae only one or two immunising exposures were used. The degree of protection in the adult worm load (9-11%) was not significant and no significant differences were noticed in the pathology of the vaccinated and control animals. In the third experiment four doses of irradiated organisms were used to vaccinate five groups of sheep: 3Kr or 6Kr cercariae were administered by percutaneous infection; 6Kr skin-transformed schistosomula were administered by intra muscular injection; the same 6Kr skin-transformed schistosomula were given by intravenous injection and 6Kr syringe transformed schistosomula were administered by intramuscular injection. The degree of protection (determined as the reduction in worm burden) achieved by these different procedures was respectively 72%, 61%, 77%, 56% and 78%. These results indicate the possibility of making a live vaccine against ovine schistosomiasis and show that effective immunisation is not dependent on the presence of a mature worm infection or on cercarial penetration of the skin by the immunising infection.     

Development of resistance to coccidiosis in the absence of merogonic development using X-irradiated Eimeria acervulina oocysts

Sporulated oocysts of the protozoan Eimeria acervulina were subjected to 0, 10, 15, 20, or 30 krad of X-irradiation and inoculated into susceptible outbred chickens to determine if radioattenuated coccidia could induce protection against parasite challenge. Irradiation treatment had an appreciable dose-dependent effect on parasite development. Insignificant numbers of oocysts were produced by chickens inoculated with parasites that had been exposed to greater than 10 krad X-irradiation. Sporozoites exposed to 15 or 20 krad irradiation conferred significant protection against the appearance of intestinal lesions after parasite challenge. Sporozoites subjected to the highest dose level (30 krad) did not produce any significant level of protection. To investigate this phenomenon further and assess intracellular parasite development, susceptible outbred strains of chickens were administered either nonirradiated (0 krad) oocysts or oocysts that were exposed to an optimal dose (15 krad) or a high dose (30 krad) of X-irradiation. Immunofluorescence staining of tissue sections from each treatment group at various intervals after the initial administration of irradiated parasites indicated that sporozoites exposed to 15 krad irradiation were as capable of invading the host intestinal epithelium as nonirradiated sporozoites. However, at 48, 60, 72, and 96 hr, there was a marked reduction in merogonic development in groups receiving irradiated sporozoites compared to those inoculated with nonirradiated parasites. The latter parasites underwent profuse merogonic development; in contrast, irradiated parasites demonstrated little (15 krad) or no (30 krad) merogonic development. These results suggest that induction of a protective immune response occurs during a critical period early in intracellular development of E. acervulina.     

Schistosoma mansoni: Vaccination of mice with highly X-irradiated cercariae

Vaccination against schistosomiasis with highly X-irradiated Schistosoma mansoni cercariae was studied in mice. The optimum dose of X radiation for the attenuation of cercariae was in the range of 24–48 krad. In selecting the optimum dose, lesions caused by migrating schistosomula in the lungs of the immunized host were considered. Cercariae exposed to 48 krad caused fewer lesions than those exposed to 24 krad but still effected a comparable worm reduction. The percentages of worm reduction in mice immunized with 48-krad X-irradiated cercariae increased with the number of immunizations up to the fifth immunization and then fluctuated in the sixth, seventh, and eighth days without increase. The optimum dose of immunizing cercariae was 500, and the optimum time interval for successive immunizations was 4 weeks. There was no significant difference in susceptibility to infection in the adult mice 161 to 694 days of age. The duration of acquired immunity in immunized mice is long, still evident 545 days from the last immunization. The present studies clearly showed that with the bioengineering method, the worm reduction in the immunized mice reached 91.1%, the effect of immunization was stronger in mice immunized with the highly X-irradiated cercariae than with the low X-irradiated cercariae, and X-irradiated cercariae were demonstrated to be a strong inducing agent for immunity in mice.     

Efficacy of UV-irradiated larval vaccine of Ancylostoma ceylanicum (Looss, 1911) in golden hamsters (Mesocricetus auratus)

A vaccination trial in golden hamsters with UV-irradiated infective larvae of Ancylostoma ceylanicum was attempted. One oral vaccination of hamsters with 100 infective larvae irradiated by means of UV-tube (390 nm) at different time intervals induced the development of resistance. As the time exposure of irradiation was increased, there was a corresponding decrease in the subsequent worm establishment. A high level of protection afforded by larvae irradiated for 15 min UV-exposure was recorded giving 99.0% and 95.0% worm reduction against the challenge doses of 100 and 1000 normal larvae respectively. There was no marked difference in worm establishment in hamsters vaccinated either orally or subcutaneously, followed by oral challenge. In the vaccinated hamsters, the manifestations of resistance at 15 min UV-exposure were shown by marked reduction in worm establishment and highly reduced epg in pellets with significantly higher blood haemoglobin levels compared with those given normal larvae as vaccine and challenge controls.     

Schistosoma mansoni: parasitology and immunology of baboons vaccinated with irradiated cryopreserved schistosomula

Young, captivity-born male baboons (Papio cynocephalus) were vaccinated with γ-irradiated (500 Gy) cryopreserved Puerto Rican strain schistosomula of S. mansoni. Protection against heterologous, normal Kenyan Strain S. mansoni challenge infection was erratic and partial; and two putative correlates of immunity, reduced worm fecundity and change in worm location (anterior shift) were not observed. However, immunization of baboons with this vaccine resulted in a stimulated immune system. Both cellular and humoral anamnesis were demonstrable in vaccinated-challenged baboons. Schistosome infection-associated IgM hypergammaglobulinema was also greatly reduced in vaccinated-challenged baboons. On the other hand, IgG antibodies to adult, egg, and cercarial antigens were increased after challenge infection in preimmunized baboons. Vaccination appears to have resulted in a redirection of the immune system into anti-parasite channels, but this more specific immune response was insufficient to confer good protection against challenge infection in this experiment. The dampening effect of the vaccine on the hypergammaglobulinemia of schistosomiasis is another candidate for a possible “anti-pathogenesis” effect of irradiated schistosome larval vaccines, to be added to the reduced granuloma size already reported (Damian, Roberts, Powell, Clark, Lewis &; Stirewalt, 1984) to occur in these vaccinated baboons. Together, they could increase the potential benefit of live, irradiated vaccines for schistosomiasis. Human trials with vaccines only partially protective against challenge infections may therefore ultimately be warranted.     

Comparison of the protective resistance induced by 60Co-irradiated cercariae and schistosomula of the WFFS and NMRI strains of Schistosoma mansoni

Mice, CBA/HT6T6 and C57BL/10, were vaccinated with 1 X 350 or 1 X 500 Schistosoma mansoni cercariae or schistosomula attenuated with 20 or 56 krad 60Co irradiation and challenged with 200 cercariae. Protective resistance against homologous strain challenge was compared using the Winches Farm Field Station (WFFS) and Naval Medical Research Institute (NMRI) strains of S. mansoni. Maximal resistance to challenge was obtained in both strains of mice with cercariae or schistosomula of either WFFS or NMRI strain attenuated with 20 krad. Protection using organisms attenuated with 56 krad was significantly lower. Since previous studies with the two parasite strains have shown that the biological effects of irradiation are similar, the difference in the immunogenicity of the 56 krad-irradiated NMRI strain in this study from earlier studies must be due either to different local conditions for irradiation or to adaptation of the NMRI strain to a new laboratory environment. This finding may have important implications for vaccination studies and investigations of the mechanisms of immunity where radiation-attenuated parasites are used.     

Schistosoma mansoni: vaccination of mice with 10-krad-irradiated, cryopreserved schistosomules

Protection against a Schistosoma mansoni cercarial challenge was evaluated in mice immunized with a vaccine composed of 10-krad-irradiated, cryopreserved schistosomules. The level of resistance induced in C57B1/6 or NMRI (CV) mice increased with the number of schistosomules injected. Up to 83% reduction in challenge worm burden was achieved when 5000 schistosomules were injected per mouse. Intramuscular injection of the vaccine was superior to subcutaneous. Multiple immunizations, up to 3 at 4-week intervals, did not increase the resistance induced by a single immunization. A high level of protection developed in as little as 2 weeks and was maintained through at least 12 weeks postimmunization. The vaccine irradiated with 10 krad from either a 60-cobalt or 137-cesium source induced equivalent levels of resistance, and no differences were found in the immunogenicity of vaccines comprised of organisms irradiated as cercariae or as 1- to 3-hr-old schistosomules. These findings are basic to the development of a cryopreserved, live vaccine against schistosomiasis of humans or domestic animals.     

Acanthocheilonema viteae: vaccination of jirds with irradiation-attenuated stage-3 larvae and with exported larval antigens

Jirds (Meriones unguiculatus) were immunized with irradiated (35 krad) stage-3 larvae (L3) of Acanthocheilonema viteae. The induced resistance against homologous challenge infection and the antibody response of the animals were studied. Immunization with 3, 2, or 1 dose of 50 irradiated L3 induced approximately 90% resistance. Immunization with a single dose of only 5 irradiated L3 resulted in 60.8% protection while immunization with a single dose of 25 L3 induced 94.1% protection. The protection induced with 3 doses of 50 irradiated L3 did not decrease significantly during a period of 6 months. Sera of a proportion, but not all resistant jirds, contained antibodies against the surface of vector derived L3 as defined by IFAT. No surface antigens of microfilariae or adult worms were recognized by the sera. Vaccinated animals had antibody responses against antigens in the inner organs of L3 and in the cuticle and reproductive organs of adult worms as shown by IFAT. Immunoblotting with SDS-PAGE-separated L3 antigens and L3-CSN revealed that all sera contained antibodies against two exported antigens of 205 and 68 kDa, and against a nonexported antigen of 18 kDa. The 205-kDa antigen easily degraded into fragments of 165, 140, 125, and 105 kDa which were recognized by resistant jird sera. Various antigens of adult worms, but relatively few antigens of microfilariae, were also recognized. To test the relevance of exported antigens of L3 to resistance, jirds were immunized with L3-CSN together with a mild adjuvant. This immunization induced 67.7% resistance against challenge infection and sera of the immunized animals recognized the 205- and 68-kDa antigens of L3.     

Dipetalonema viteae: resistance in Meriones unguiculatus with multiple infections of stage-3 larvae

The jird, Meriones unguiculatus, infected with 80 normal infective larvae of Dipetalonema viteae, revealed a recovery rate of 27.9% 12 weeks after infection. A pretreatment by three injections of 50 normal larvae each and challenge by 80 larvae resulted in a recovery rate of 10.7%. The recovered worms were longer than those from the challenge control animals. When three times 50 irradiated larvae (35 krad) were inoculated, the recovery rate of the challenge decreased to 2.6%, representing a protection of 90.7%. The surviving adult worms were stunted and derived exclusively from the 80 normal larvae given for challenge, since absolutely no adult worms were recovered in eight animals inoculated three times with 50 irradiated larvae only. Sera of all pretreated jirds contained IgG and IgM antibodies which bound in immunoblotting experiments bound predominantly to three proteins of larvae with molecular masses of 68,140, and 165 kDa, respectively. Enzymatic surface iodination revealed that the three antigens were exposed on the larval surface. The coincidence of a partial resistance to a challenge infection and of an antibody response against surface proteins of infective larvae suggests an importance of these antigens for the rejection of D. viteae mediated by an acquired immunological resistance of M. unguiculatus.     

The ability of fractionated sera from animals vaccinated with irradiated cercariae of Schistosoma mansoni to transfer immunity to mice

To study the role of IgG and IgM isotypes found in sera of mice or rabbits immunized with irradiated cercariae in schistosome immunity, the respective sera were fractionated by protein A chromatography. Both the protein A-bound and unbound fractions of vaccinated mouse serum (VMS) showed reactivities in ELISA assay using NP-40 membrane extracts of 3-hr schistosomula as antigens and in indirect immunofluorescence assay (IIF) using live 3-hr schistosomula. Both the protein A-bound and unbound fractions possessed high levels (84% and 76%, respectively) of complement-mediated cytotoxicity against schistosomula in vitro. The IgG- and IgM-containing fractions each conferred passive protection (30% and 20%) against challenge infection, although at a lower level when compared to unfractionated VMS (42%). These data demonstrate that in the mouse model both IgG and IgM can recognize epitopes on the surface of schistosomula, mediate cytotoxicity in vitro, and provide passive protection in vivo. Similarly, the protein A-bound and unbound fractions of vaccinated rabbit serum (VRS) were also shown to be positive in ELISA and IIF. The IgG- and IgM-containing fractions each possessed high levels (95% and 85%, respectively) of complement-dependent cytotoxicity against schistosomula in vitro. In contrast to VMS fractions, the IgG fraction of VRS conferred a similar level (28%) of in vivo protection as unfractionated VRS when injected into mice no later than 6 days after challenge. Moreover, the IgG fraction of VRS was still able to provide passive protection to mice when given as late as 15 days postinfection, but failed to confer protection when injected at 24 or 35 days postinfection.(ABSTRACT TRUNCATED AT 250 WORDS)     

Effects of gamma radiation on spermatophore production and reception and subsequent fecundity and egg viability inAcarus siro L. (Acari: Acaridae)

Gamma radiation at doses higher than 10 krad significantly lowered the fecundity of the grain mite,Acarus siro. The fecundity of irradiated females was inversely correlated with dose, both when control or irradiated males were used in the pairing.Irradiation with ionizing radiation affected sexual activity of males. At doses above 10 krad the number of formed or observed spermatophores was lowered significantly.Sexual attractiveness of irradiated and control females to irradiated males was similar. However, non-irradiated males were observed mating more often with non-irradiated than with irradiated females.No correlation was found between the numbers of spermatophores present in the spermathecae of the female and the fecundity of the female. Irradiation had a greater effect on fecundity of the female than on sexual activity of the male; it did not affect the shape or behavior of spermatophores in the spermathecae of the female.Viability of eggs laid by females decreased by at least 50% when females or males were irradiated with doses above 20 krad. Irradiation also affected the survival of adults, but females were more sensitive than males. Net sterility index indicates that doses higher than 20 krad induce more than 90% sterility.     

Protective monoclonal antibodies from mice vaccinated or chronically infected with Schistosoma mansoni that recognize the same antigens

An IgM monoclonal antibody, designated mAb 1.G1, has been generated from spleen cells of mice immunized with irradiated Schistosoma mansoni cercariae. As determined by indirect immunofluorescence, mAb 1.G1 binds to the surface membrane of schistosomula and to the ciliated plates of miracidia. mAb 1.G1 also binds to the protonephridial systems of live adult worms and denuded, acetone-fixed schistosomula. Western blot analysis shows that the target epitope of this mAb is found on Nonidet P-40-solubilized schistosomular antigens ranging in molecular size from 85 to 130 kDa and ciliated plate antigens of miracidia at 92, 95, and 102 kDa. The recognized epitope in an 8 M urea adult worm extract is found on a 97-kDa molecule. In addition, mAb 1.G1 mediates a high level of complement-dependent cytotoxic activity against schistosomula when used in an in vitro assay. In passive immunization experiments, approximately 40% protection was provided mice when mAb 1.G1 was administered either at the time of challenge or when given 8 days postchallenge. However, when administered 15 days postchallenge, mAb 1.G1 failed to mediate passive protection. The ability of mAb 1.G1 to mediate protection in vivo correlates with its recognition of epitopes on the surfaces of live schistosomula up to 8 days but not at 15 days. Western blot analysis showed that the antigens were contained within Nonidet P-40 extracts of schistosomula during the same time period. Furthermore, a second monoclonal antibody (mAb 4.4B) derived from mice chronically infected with S. mansoni exhibits the identical properties as described for mAb 1.G1.     

Effect of Gamma-irradiation on Growth, RNA, Protein, and Nitrogen Contents of Bean Callus Cultures

Callus-tissue cultures of Phaseolus vulgaris L. were subjectedto various doses of 60Co gamma-irradiation, and its effect ongrowth, total RNA, soluble protein, and nitrogen contents hasbeen studied. The growth of the tissue cultures was stimulatedby low levels of radiation (0.5 krad). However, from 1 to 10krad, there was a gradual and linear decrease in growth. Thecells exhibited a wide variety of shapes and sizes, mitoticinhibition, degeneration of cytoplasm, browning of the cellwall, and reduced plating efficiency. At 20–30 krad growthwas drastically reduced, followed by severe killing of the cellsand cessation of growth at 40 krad. With increase in dosimetry,RNA, and soluble protein continued to decrease. At lower doses(0.5 and 1 krad) there was no significant difference in totalnitrogen of the control and irradiated cultures, however, from2 krad upwards there was a gradual increase in total nitrogenin terms of µg/mg dry weight of the irradiated callus.The results demonstrate that there is a direct correlation betweengrowth, RNA and protein levels. Gamma-irradiation in generalcaused inhibition of tissue culture growth along with failureof RNA, and subsequently of protein synthesis.     

Babesia bigemina: immune response of cattle inoculated with irradiated parasites

Effects of various radiation dosages on the infectivity and immunogenicity of Babesia bigemina were studied. Calves infected with 1 × 10¹⁰, B. bigemina parasitized erythrocytes exposed to 24 krad developed progressive parasitemias. Some calves receiving 1 × 10¹⁰ parasitized erythrocytes irradiated at 36 krad did not develop progressive infections. Progressive infections were prevented by exposure to irradiation at 48 and 60 krad. A degree of acquired resistance to infection with B. bigemina developed in calves after inoculation with parasites irradiated at 48 and 60 krad. The resistance developed was sufficient to suppress multiplication of the Babesia and to permit calves to survive otherwise severe clinical infections due to challenge with nonirradiated parasites. Irradiated parasites were frozen without apparent loss of immunizing properties.     

Density-dependent effects on the survival and growth of the rodent stomach worm Protospirura muricola in laboratory mice

The spirurid nematode, Protospirura muricola, is of intrinsic interest as a rodent model of gastric nematode infections. Since worm burdens can be very heavy in nature, density dependent processes may constrain parasite growth. Laboratory mice (BKW) were exposed to varying doses of infective larvae of P. muricola in the range 5 to 40 third-stage larvae (L3), in four separate experiments in which progressively higher doses were utilized. All mice were culled 60 days after infection and a total of 518 worms (226 male and 292 female worms) was recovered, measured and weighed. Overall survival was 58.9%, but survival declined significantly with increasing dose by approximately 21% (from 66% at 5 L3 per mouse to 52% at 40 L3 per mouse). The length and weight of worms correlated positively in both sexes. Total worm biomass increased linearly with increasing numbers of worms. However, whilst the length and weight of male worms declined with increasing worm burden (8.4 and 24.6% respectively), female worms were less affected, only length showing a significant reduction with increasing parasite burden (16.0%). Therefore, increasing worm burdens impeded growth of P. muricola, but reduction in length and weight were relatively small in relation to the overall size of this nematode. Increasing worm burdens were associated with loss of host weight and reduction in stomach weight and worm burdens in excess of 20 exerted a measurable cost to the host, which in the field, may be associated with loss of overall host fitness.