Determination of Giardia lamblia cyst infective dose for the Mongolian gerbil (Meriones unguiculatus).

The purpose of this study was to determine the 50% infective dose for Giardia lamblia (CDC:0284:1) cysts in Mongolian gerbils (Meriones unguiculatus). The log10 50% infective dose results calculated by probit analysis and the Spearman-Karber method were 2.45 and 2.50, respectively.     

Efficient Colonization of the Bean Bug Riptortus pedestris by an Environmentally Transmitted Burkholderia Symbiont

The bean bug Riptortus pedestris is specifically associated with the Burkholderia gut symbiont and acquires the symbiont from the environment every generation. Here, we investigated the infective dose of the symbiont by experimental administration. The 50% infective dose was remarkably low, only 80 cells, indicating efficient colonization of the symbiont.     

Dose response of Cryptosporidium parvum in outbred neonatal CD-1 mice.

Cryptosporidium parvum infectivity in a neonatal CD-1 mouse model was used to determine the dose needed to infect 50% of the population. The 50% infective dose was estimated to be 79 oocysts. It was observed that a mean oral inoculum of 23 oocysts produced infection in 2 of 25 neonatal mice 7 days postinoculation. All animals became infected when the mean oral dose exceeded 310 oocysts per animal. The dose response of C. parvum was modeled with a logit dose-response model suitable for use in water disinfection studies.     

The preservation of infective spores of Octosporea muscaedomesticae in Phormia regina,of Nosema algerae in Anopheles stephensi,and of Nosema whitei in Tribolium castaneum by lyophilization

Infective spores of three species of microsporidia were subjected to the lyophilization process by employing varying media as cryoprotectants. The infectivity of the lyophilized spores was then tested against a standard fresh spore preparation in the appropriate host insect. Spores of Octosporea muscaedomesticae served as an experimental model and were rendered noninfective in host Phormia regina (Calliphoridae: Diptera) after lyophilization with the following cryoprotective agents: skim milk (12%), ascorbic acid (5%) combined with thiourea (5%), glycerol (10%), mesoinositol (5%), and equine serum. Spores of O. muscaedomesticae lyophilized or vacuum-dried in 50% sucrose as well as in the hosts' tissues remained highly infective for as long as 2 years at a dose of 10⁶ spores/fly and a trial length of 12 days. At a dose of 5 × 10⁴ spores/fly there was a slight decrease in infectivity of the spores which had been lyophilized in the host's abdomen after a 2-year storage period compared with that of fresh, nonlyophilized spores. Naked spores of Nosema algerae suspended in 50% sucrose and lyophilized produced infection in 50% of the host population of Anopheles stephensi (Culicidae: Diptera) compared with 70% infection produced by fresh non-lyophilized spores. Spores of Nosema whitei lyophilized within its host larva Tribolium castaneum (Tenebrionidae: Coleoptera) remained 100% infective at a dose of 5 × 10⁵ spores/gram diet. It is concluded that an aqueous solution of 50% sucrose and/or the host's tissues are excellent protectants for the cryogenic or vacuum-drying process of the above-named spores, and their protective function may apply also to other microsporidian species.     

Intracutaneous infection with Leptospira icterohaemorrhagiae (Shibaura strain) of the guinea pig

Experimental leptospirosis with Leptospira icterohaemorrhagiae Shibaura strain was studied in guinea pigs. When the pathogen was inoculated intracutaneously to the back of the animals, localized haemorrhage was observed at the inoculated site before the appearance of general haemorrhage. The severity of the local lesion increased progressively until the 7th day of inoculation. The minimum infective dose (MID) or the 50% infective dose (ID50) of the leptospiral suspension was determined by the appearance of the macroscopic local haemorrhage 7 days after inoculation. The MID thus determined was almost comparable with the value determined by the development of general symptoms and signs by conventional ip inoculation. The number of the pathogen per ID50 varied between 6 and 35 in five experiments. The local haemorrhage was effectively protected by active or passive immunization. Microscopically, haemorrhage at the inoculated site was found mainly in the dermis, directly beneath the epidermis in particular, and accompanied with leakage of the pathogen. The pathogen was also detected abunduntly in the thickened epidermal layer covering the inoculated area as well as in the epithelial matrix of hair-follicle, probably due to the proliferation of the pathogen at the site.     

The fate of multiple doses of infective larvae of Nematodirus battus in 8-month-old lambs and their effect on intestinal enzyme activity

Mapes C.J. and Coop R.L. 1973. The fate of multiple doses of infective larvae of Nematodirus battus in 8-month-old lambs and their effect on intestinal enzyme activity. International Journal for Parasitology3: 363–370. The fate of five daily doses of 60,000 infective larvae of Nematodirus battus was studied in 8-month-old lambs. On 18, 26 and 34 days after the last larval dose 4.0, 3.3 and 1.0 per cent of the total infective dose was recovered. Approximately 50 per cent of the worms recovered on these days were fourth-stage larvae. It is suggested that L5 and adult stages were preferentially lost from the hosts and that male worms were developing at a faster rate than the females. The populations of N. battus were smaller, contained higher proportions of fourth-stage larvae and shorter L5 and adult worms than those developing from similar infective doses in 3-month-old lambs. A transient decrease in alkaline phosphatase and maltase levels was found in the mucosa of the small intestine and was compared with the marked and persistent changes in mucosal enzyme activities found with similar infective doses in 3-month-old hosts.     

Infectivity and reproductive potential of the Oswego strain of Heterorhabditis bacteriophora associated with life stages of the clover root curculio,Sitona hispidulus

The infectivity and reproductive potential of the entomopathogenic nematode Heterorhabditis bacteriophora (Oswego strain), at different concentrations, was studied. Seventy to 80.0% mortality to late instar larvae of the clover root curculio, Sitona hispidulus, and 40.0-76.0% mortality to pupae, was observed at concentrations of 15-100 infective juveniles. There were no significant differences in mortality among nematode concentrations. LC(50) levels of 4.0 and 21.4 nematodes were determined for clover root curculio larvae and pupae, respectively. Nematodes did not cause significant mortality to adult or first instar clover root curculio. H. bacteriophora was able to complete its development and reproduce in 74.0-95.0% of clover root curculio late instar larvae and pupae. Reproductive potential in curculio larvae and pupae ranged from 0 to 7040 infective juveniles per host. Larvae exposed to 100 nematodes had a reproductive potential significantly higher than in those larvae exposed to 15 and 50 nematodes. Reproductive potential in pupae decreased with an increased nematode dose, indicating potential crowding effects. Host larval and pupal mass were positively correlated with nematode progeny production.     

Susceptibility of the peachtree borer, Synanthedon exitiosa, to Steinernema carpocapsae and Steinernema riobrave in laboratory and field trials

The nematode Steinernema carpocapsae (All) strain was significantly more effective against peachtree borer larvae (Synanthedon exitiosa [Lepidoptera: Sesiidae]) than Steinernema riobrave (7-12) strain in field and laboratory experiments. Eighty-eight percent control of peachtree borer larvae was obtained with S. carpocapsae in the field trial when applied at 3 x 10(5) infective juveniles per tree, and 92% mortality was obtained in the lab assay using 50 infective juveniles per larva.     

Comparison of the infectivity of Trypanosoma cruzi insect-derived metacyclic trypomastigotes after mucosal and cutaneous contaminative challenges

Trypanosoma cruzi infects humans when infected triatomine vector excreta contaminate breaks in skin or mucosal surfaces. T. cruzi insect-derived metacyclic trypomastigotes (IMT) invade through gastric mucosa after oral challenges without any visible inflammatory changes, while cutaneous and conjunctival infections result in obvious local physical signs. In this study we compared the infectivity of T. cruzi IMT in mice after cutaneous and oral contaminative challenges simulating natural infections. The 50% infective dose (ID50) for oral challenge was 100 fold lower than the ID50for cutaneous challenge, indicating that oral mucosal transmission is more efficient than cutaneous transmission.     

Use of real-time PCR to detect canine parvovirus in feces of free-ranging wolves

Using real-time PCR, we tested 15 wolf (Canis lupus) feces from the Superior National Forest (SNF), Minnesota, USA, and 191 from Yellowstone National Park (YNP), USA, collected during summer and 13 during winter for canine parvovirus (CPV)-2 DNA. We also tested 20 dog feces for CPV-2 DNA. The PCR assay was 100% sensitive and specific with a minimum detection threshold of 10(4) 50% tissue culture infective dose. Virus was detected in two winter specimens but none of the summer specimens. We suggest applying the technique more broadly especially with winter feces.     

Infectivity of two nematode parasites, Camallanus lacustris and Anguillicola crassus, in a paratenic host, the three-spined stickleback Gasterosteus aculeatus

Three-spined sticklebacks Gasterosteus aculeatus are frequent paratenic hosts of the nematode parasites Anguillicola crassus and Camallanus lacustris. As paratenic hosts, sticklebacks could spread infection by carrying high numbers of infective stages. In contrast, low infective ability of either parasite for the paratenic host could hinder the spread of infection. In the present study, G. aculeatus was, for the first time, infected under controlled laboratory conditions with defined doses of the parasites. Sticklebacks were exposed to 6, 12, 18 and 24 parasite larvae to determine the infective ability of the 2 nematode species. There were significantly higher infection rates for C. lacustris (18 to 49%) than for A. crassus (4 to 14%) at each exposure dose. In C. lacustris-infected sticklebacks, infection rates tended to be highest after exposure to 12 C. lacustris larvae and lowest after exposure to 24 parasites. In A. crassus-infected sticklebacks, no effect of parasite exposure dose on infection rates was observed. Immunity parameters such as respiratory burst activity and lymphocyte proliferation of head kidney leukocytes recorded 18 wk post exposure were not significantly affected by either parasite or exposure dose. Granulocyte:lymphocyte ratios were elevated only within the stickleback group showing the highest infection intensity of C. lacustris, i.e. to those exposed 18 parasites.     

Development and validation of an egg-based potency assay for a trivalent live attenuated influenza vaccine

Live attenuated influenza vaccines (LAIVs) targeting seasonal influenza are produced in embryonated eggs and formulated as a trivalent preparation of three live attenuated vaccine strains, one A(H1N1) strain, one A(H3N2) strain and one type B strain. In this study, we describe an egg-based potency assay for estimating the 50% Egg infectious dose (EID50) of individual strains in the trivalent preparation by selective neutralisation of two strains and then estimating the infective titres of the non-neutralised strain. The test is highly specific, and no cross interference of heterologous antisera is observed in the estimation of individual titres. Individual strains with titres in the range of 6.5-7.0 log EID50 per 0.5 ml show intra-assay and inter-assay coefficients of variance ranging from 1.25% to 2.95%. This assay was developed to establish a simple, reliable and inexpensive egg-based assay for estimating the potency of individual strains in a trivalent preparation.     

A spore counting method and cell culture model for chlorine disinfection studies of Encephalitozoon syn. Septata intestinalis

The microsporidia have recently been recognized as a group of pathogens that have potential for waterborne transmission; however, little is known about the effects of routine disinfection on microsporidian spore viability. In this study, in vitro growth of Encephalitozoon syn. Septata intestinalis, a microsporidium found in the human gut, was used as a model to assess the effect of chlorine on the infectivity and viability of microsporidian spores. Spore inoculum concentrations were determined by using spectrophotometric measurements (percent transmittance at 625 nm) and by traditional hemacytometer counting. To determine quantitative dose-response data for spore infectivity, we optimized a rabbit kidney cell culture system in 24-well plates, which facilitated calculation of a 50% tissue culture infective dose (TCID(50)) and a minimal infective dose (MID) for E. intestinalis. The TCID(50) is a quantitative measure of infectivity and growth and is the number of organisms that must be present to infect 50% of the cell culture wells tested. The MID is as a measure of a system's permissiveness to infection and a measure of spore infectivity. A standardized MID and a standardized TCID(50) have not been reported previously for any microsporidian species. Both types of doses are reported in this paper, and the values were used to evaluate the effects of chlorine disinfection on the in vitro growth of microsporidia. Spores were treated with chlorine at concentrations of 0, 1, 2, 5, and 10 mg/liter. The exposure times ranged from 0 to 80 min at 25 degrees C and pH 7. MID data for E. intestinalis were compared before and after chlorine disinfection. A 3-log reduction (99.9% inhibition) in the E. intestinalis MID was observed at a chlorine concentration of 2 mg/liter after a minimum exposure time of 16 min. The log(10) reduction results based on percent transmittance-derived spore counts were equivalent to the results based on hemacytometer-derived spore counts. Our data suggest that chlorine treatment may be an effective water treatment for E. intestinalis and that spectrophotometric methods may be substituted for labor-intensive hemacytometer methods when spores are counted in laboratory-based chlorine disinfection studies.     

The multiplication and spread of sacbrood virus of bees

Sacbrood virus multiplied without causing symptoms in adult bees when it was injected into them or when it was fed to young individuals. More virus accumulated in heads of infected bees than elsewhere in their bodies, and much was in their hypopharyngeal glands. The extract of each infected head contained about 10²medial lethal doses (LD 50s) of sacbrood virus for larvae when given in their food. The infective dose of sacbrood virus by injection for adults was about 10^-4of the LD 50 in food for larvae. The infective dose by mouth for adults was about 10²LD 50s for larvae, but bees older than 4–8 days could not be infected this way. Infection did not spread between adults but is probably transmitted in nature from infected adults to larvae and back to young adults that ingest remains of larvae killed by sacbrood. The youngest adult bees infected with sacbrood ate little pollen and lived only about 3 weeks in the laboratory, as did bees receiving no pollen, whereas bees fed with ample pollen lived 9 weeks.     

Eimeria stiedae: weight, oocyst output, and hepatic function of rabbits with graded infections

Groups of 5 (38–40 days old) Eimeria stiedae-naive rabbits were infected with 0 (mock infection), 10², 10³, 10⁴, and 10⁵ sporulated oocysts of Eimeria stiedae (groups A, B, C, D, and E, respectively) and body weight, oocyst output, serum glutamic pyruvic and serum oxalacetic transaminases, bilirubinemia. lipemia, glycemia, and proteinemia were measured on diverse occasions for 50 days. Mortality and carcass and liver weights at the end of the experiment were also recorded. Mortality was 80% in group E, 40% in group D, and 0% in the other groups. Reduction of weight gain was observed from the 8th day of infection and actual loss from the 15th day in infected animals. On Day 50, the average body and carcass weights of all infected rabbits were 71.2 and 63.2%, respectively, of group A. Only groups D and E had absolute hepatomegaly and group C had relative liver enlargement. Patency and rate of increase of oocyst output were not related to dose but peak and declination of oocysts production were delayed in proportion to the infective dose.Serum glutamic pyruvic and glutamic oxalacetic transaminases were increased from Day 8 to Day 36, bilirubinemia and lipemia augmented from Day 22, and glycemia and total serum proteins decreased from Days 22 and 29, respectively. Bilirubinemia tended to recover sooner (toward Day 50) in rabbits with lighter infection and lipemia recovered from Day 36 in proportion to the size of the infective dose. Modifications of glycemia and total proteinemia were consistent but reached statistical significance only occasionally. The asexual reproduction of the parasites caused transient damage to the hepatocytes during the second week of infection, and later sexual stages altered the ductal epithelium from the fourth week.     

The fate of large infective doses of Nematodirus battus in young lambs

Mapes C.J., Coop R.L. and Angus K.W. 1973. The fate of large infective doses of Nematodirus battus in young lambs. International Journal for Parasitology3: 339–347. The fate of 300,000 and 5 × 60,000 infective larvae of Nematodirus battus during the 2–3 weeks after patency was studied in 3-month-old lambs. The proportions of the infective doses recovered after 18, 26 and 34 days were 37, 6 and 3.5 per cent in the single dose and 29,12 and 4.7 in the multiple dose groups. Fifth-stage larvae and adults, especially females, were preferentially lost from the hosts. The numbers of worms present and the percentage of fourth-stage larvae present were similar in both the single and multiple dose groups.     

Necator americanus: temperature, pH, light, and larval development, longevity, and desiccation tolerance

The effect of incubation temperature and pH on the hatch rate of eggs of Necator americanus, and the desiccation tolerance of the resulting infective stage-3 larvae were investigated in the laboratory under controlled conditions. Hatching did not occur below 15 C and above 35 C. A 21% hatch rate was obtained at 15 C while a 10.6% hatch rate was obtained at 35 C. The highest hatch rate (93.7%) was obtained at 30 C. The optimum pH for hatching was 6.0, but the larvae did not reach the infective stage. Incubation temperature of the eggs affected the longevity and desiccation tolerance of resultant infective larvae. Larvae hatched at 30 C and maintained at 26 C under bright fluorescent light had a 50% survival time (S50) of 4 days. In the dark or shade, the S50 for larvae raised at 30 C was 5 weeks, while that of larvae hatched at 20 C was 7 weeks. Incubation temperature also affected the desiccation tolerance of larvae. Larvae developed at 20 C were more resistant to desiccation at various relative humidity values than larvae hatched at 30 C.     

Cellular immune response to hog cholera virus (HCV): T cells of immune pigs proliferate in vitro upon stimulation with live HCV, but the E1 envelope glycoprotein is not a major T-cell antigen.

T-cell responses of pigs to hog cholera virus (HCV) have reportedly been absent or difficult to detect. Therefore, little is known about cellular immunity to HCV. In this study, we used an attenuated strain of pseudorabies virus expressing the envelope glycoprotein E1 of HCV and purified recombinant E1 to examine whether the E1 protein is a target antigen recognized by the T cells of HCV-immune pigs. We were unable to identify the E1 protein as a major target antigen recognized by the T cells of HCV-immune animals. However, such cells proliferated in vitro upon stimulation with viable HCV antigen. The lymphoproliferative response to HCV was strictly time and dose dependent and could be induced upon stimulation by live but not by UV light-inactivated HCV. Depletion studies demonstrated that lymphoproliferation depended on the presence of CD2+CD8bright+ lymphocytes, but CD2+CD4+ cells also contributed to the lymphoproliferative response. The primary lymphoproliferative response in animals inoculated with 10(7) 50% tissue culture infective doses of strain Brescia 2.1.1 was stronger than that observed in animals inoculated with 10(3) 50% tissue culture infective doses of the Cedipest strain. A remarkable finding was the increase in non-antigen-specific lymphoproliferation upon inoculation of the animals with HCV strains. This immunological phenomenon may mask a specific T-cell response to the virus.     

Mechanisms of Pathogenesis, Infective Dose and Virulence in Human Parasites

The number of pathogens that are required to infect a host, termed infective dose, varies dramatically across pathogen species. It has recently been predicted that infective dose will depend upon the mode of action of the molecules that pathogens use to facilitate their infection. Specifically, pathogens which use locally acting molecules will require a lower infective dose than pathogens that use distantly acting molecules. Furthermore, it has also been predicted that pathogens with distantly acting immune modulators may be more virulent because they have a large number of cells in the inoculums, which will cause more harm to host cells. We formally test these predictions for the first time using data on 43 different human pathogens from a range of taxonomic groups with diverse life-histories. We found that pathogens using local action do have lower infective doses, but are not less virulent than those using distant action. Instead, we found that virulence was negatively correlated with infective dose, and higher in pathogens infecting wounded skin, compared with those ingested or inhaled. More generally, our results show that broad-scale comparative analyses can explain variation in parasite traits such as infective dose and virulence, whilst highlighting the importance of mechanistic details.     

J亚群禽白血病病毒NX0101株的TCID50与p27抗原之间的相关性研究

为了研究J亚群禽白血病病毒在细胞上接种后的半数细胞培养物感染量(TCID₅₀)与p27抗原的酶联免疫吸附试验（ELISA）的S/P值之间的相关性,并探讨其意义。本试验将J亚群禽白血病病毒NX0101株接种鸡胚成纤维细胞（CEF细胞）,换维持液后连续10d取样,检测10d的TCID₅₀值与p27抗原的S/P值之间的相关性。同时,将该毒株在DF-1细胞系上传代至20代,取其中的第1代、第5代、第10代、第15代和第20代分别进行TCID₅₀滴度的测定和p27抗原检测。结果表明:在CEF细胞上接种的NX0101株J亚群禽白血病病毒连续10d的TCID₅₀值与p27抗原之间存在显著的相关性（r=0.85277;P<0.0001）呈正相关;在DF-1细胞系上传的不同代数之间也呈显著正相关（r=0.93000;P=0.0220）。由此可以推测J亚群禽白血病病毒的TCID₅₀与p27抗原呈显著正相关,因而可以用ELISA法测得的p27抗原的S/P值对病毒的TCID₅₀值进行估测。