Nonenzymatic isolation of Trichinella pseudospiralis infective L1 larvae at pH 7.4

Over half of the number of Trichinella pseudospiralis infective L1 larvae recovered from host carcasses by pepsin-HCl digestion were isolated from homogenized carcasses incubated in HBSS. More worms isolated by the latter method were viable compared to those isolated by pepsin-HCl digestion. When host carcasses infected with T. pseudospiralis were diced into pieces and incubated in HBSS, 30% more worms were recovered than from homogenized carcasses incubated in HBSS as above, and the majority of worms acquired by the former method were viable. The infectivity of T. pseudospiralis infective L1 larvae isolated from homogenized muscle in HBSS was 3.9 times greater than that for larvae recovered from homogenized carcasses by pepsin-HCl digestion. Only 4% and 0.8% of the number of T. spiralis recovered from homogenized muscle by pepsin-HCl digestion were isolated from homogenized or diced muscle incubated in HBSS, respectively. Fewer T. spiralis isolated from homogenized tissue in HBSS were viable compared to those recovered from homogenized carcasses digested in pepsin-HCl or diced carcasses incubated in HBSS.     

Characterization of Nippostrongylus brasiliensis infection in different strains of mice

Five mouse strains, CBA/J, BALB/c, C3H/HeJ, A/J, and C57Bl/6J-bg-bg, all showed similar expulsion kinetics for Nippostrongylus brasiliensis (infective dose = 500 L3). Typically, parasite recovery was maximal on day 2 in the lungs and by day 4 in the small intestine. Few worms (less than 5% infective dose) were recovered on day 14 in all strains. These same mouse strains exhibited immune depression on day 5 of infection with mesenteric lymph node cells (MLN) showing reduced (10-30% normal) IgM, IgG, and IgA responses against heterologous antigen. The intestinal mast cell numbers and tissue histamine levels were examined in CBA/J mice. Mast cell numbers increased (normal = less than 1/villous crypt unit; VCU) from day 5 and peaked on day 12 (greater than 15/VCU). Intestinal histamine levels did not completely correlate with mast cell numbers with maximum concentrations (240 +/- 73 ng/g, 2-fold over normal) reached by day 8. Histamine concentrations in the intestine returned to normal levels by day 20.     

Infectivity of Echinococcus granulosus protoscolices under different conditions of temperature and humidity

The aim of this study was to examine the effect of different temperatures and humidities on the infectivity of Echinococcus granulosus protoscolices. Eighteen dogs (6 groups, n = 3 each) were fed with offal mince harbouring approximately 20,000 protoscolices of E. granulosus of different viabilities. Dogs were infected with E. granulosus protoscolices of: (1) 5% viability at -10 degrees C and 50% relative humidity (RH); (2) 30% viability at 0 degrees C and 60% RH; (3) 20% viability at +10 degrees C and 65% RH; (4) 15% viability at +30 degrees C and 75% RH; (5) 11% viability at +40 degrees C and 80% RH; (6) 68% viability (control group). Dogs in each group were necropsied at 29-49 days post-infection. Mean intensities of E. granulosus recovered from dogs were 256.7 +/- 60.3 in the second group; 32.7 +/- 7.1 in the third group; 40.3 +/- 15.5 in the fourth group and 1533 +/- 513 in the control group. However, no parasites were recovered from the first and fifth groups. Results obtained in the present study show that larval stages could be infective for 1 to 4 weeks during spring, autumn or winter months when maximal temperatures are approximately 0-10 degrees C. In conclusion, cold-storage depots in slaughterhouses and abattoirs where sheep carcasses might be discarded should be kept at -20 degrees C for 2-3 days, dogs should be properly controlled and adequate control programmes must be established in areas where the disease is endemic.     

Possible direct effect of diethylcarbamazine on the infective larvae of Brugia pahangi

The direct action of diethylcarbamazine (DEC) on the infective larvae of Brugia pahangi was studied. The larvae were cultured in RPMI 1640 supplemented with foetal bovine serum and antibiotics for 22 days. Most of the larvae remained alive for 8 days, but survival rate of larvae decreased rapidly from day 10 onwards. The larvae did not grow in the culture system. The addition of DEC did not affect the morbidity of the larvae and no difference was observed in the morphological characteristics between the larvae cultured in the presence or absence of DEC. The infective larvae were cultured in vitro for 5 days in the presence or absence of DEC, and inoculated into jirds. The animals were necropsied at intervals, and developing larvae and adult worms were recovered. When the larvae were cultured without DEC and then inoculated subcutaneously into jirds, 29.8% of the inoculum was recovered 3-15 days, and 25% 19-22 weeks, post-inoculation. However, when the larvae were exposed to DEC in vitro and inoculated into jirds, the rate of recovery was reduced to 25% 3-15 days post-inoculation and 2% after 19-22 weeks. When the control larvae cultured in vitro were inoculated intraperitoneally into jirds, 41.3% of inoculum was recovered 3-15 days, and 42.8% 19-22 weeks, post-inoculation. Again the corresponding value for larvae exposed to DEC in vitro was reduced to 19.8% 3-15 days, and 8% 19-22 weeks, post-inoculation. It was observed that the larvae exposed to DEC in vitro were retarded in their development in jirds.(ABSTRACT TRUNCATED AT 250 WORDS)     

Dracunculus insignis: experimental infection in the ferret, Mustela putorius furo

The laboratory study of dracunculiasis has suffered from the lack of a suitable, readily available animal model. We have been able to experimentally infect ferrets. Mustela putorius furo, with the North American dracunculid, Dracunculus insignis. Ferrets were inoculated with 75 to 100 infective larvae and were necropsied 90 to 240 days later. Guinea worms were recovered from 10 (56%) of 18 ferrets. A total of 44 worms were recovered, for an average of 4.4 worms per infected ferret. Gravid female worms were recovered as early as 128 days postinoculation. Thirteen (87%) of 15 gravid female worms were recovered from the extremities. Living male worms were recovered at 200 days of age, indicating that not all male worms die shortly after mating. First-stage larvae recovered from gravid females as early as 200 days of age were found to be infective to the copepod, Acanthocyclops vernalis. These observations suggest that the ferret is an excellent laboratory animal for dracunculiasis research.     

Stage-specific antigens of Nematospiroides dubius Baylis, 1926 (Nematoda: Heligmosomides)

Antigens were identified from Nematospiroides dubius recovered from outbred Quackenbush mice between 4 and 10 days postinoculation (PI). Parasite surface proteins were radioiodinated and extracts of the whole worms were separated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, transferred to nitrocellulose, and reacted with normal and immune mouse sera followed by an avidin-biotin-peroxidase assay. Antigens ranged between 250,000 and 20,000 molecular weight (MW). A major surface antigen, 60,000 MW, which appeared to be a complex of different antigens, and a 250,000-MW internal antigen were found on fourth-stage (L4) and fifth-stage (L5) larvae 5-10 days PI but not earlier. A group of minor surface antigens (24,000-30,000 MW) were also expressed as larvae molted from L4 to L5, 6 and 7 days PI, but they differed from antigens of similar MW expressed by adult worms. An antigen, 45,000 MW, was detected in worms 5-10 days PI, but it was only expressed on the surface of L5 worms 9 and 10 days PI. We suggest that the antigen(s) common to adults and larvae may account for protective immunity.     

Apodemus sylvaticus, a new host for Acanthocheilonema viteae (Nematoda: filarioidea)

Susceptibility of Apodemus sylvaticus and A. agrarius to infection with Acanthocheilonema viteae was compared with that of hamsters and jirds. Microfilaremia in A. sylvaticus was first noted on day 52 post-infection (p.i.) and lasted during the course of the study (up to day 150 p.i.). Maximum microfilaremic levels (female worm basis) of A. sylvaticus [mean +/- S.D. (n) = 690 +/- 1288(6)] were considerably higher than those of hamsters [16 +/- 18(6)] and jirds [51 +/- 25(5)]. Adult worm recovery in A. sylvaticus ranged from 2 to 40% of the number of infective larvae inoculated. Worm development in A. sylvaticus resembled that in hamsters and jirds. In contrast, microfilaremia was not detected in, nor adult worms recovered from A. agrarius throughout the study.     

Trichinella spiralis: quantitative relationships between intestinal worm burden, worm rejection, and the measurement of intestinal immunity in inbred mice

The effect of widely different doses of Trichinella spiralis muscle larvae on time to rejection of intestinal adults and on host survival was assessed in mice of the three rejection phenotypes; strong, intermediate, and weak. Rejection is weak with doses of less than 50 larvae per mouse. At these doses all mice rejected worms at a similar rate and no phenotypic variation was evident among strains. In contrast, rejection time was shortest for all strains and phenotypic variation among strains was evident in the range 50-100 muscle larvae/mouse. Above this dose the time taken to rejection increases monotonically with dose for all mouse strains examined. Despite this, the relative strength of rejection (i.e., phenotype) of a given strain of mouse was not changed at higher doses. Based on an end point of 98% rejection of the infective dose, time to rejection was predictable to +/- 1 day for all mouse strains and doses tested over the range 100-1000 worms administered. The principal reason for the increased time to complete rejection with larger worm doses was a delay in the initiation of intestinal rejection. This delay was evident above a dose of 50-100 larvae per mouse and occurred in all strains. Once begun, rejection was faster and eliminated more worms in unit time at higher doses (400-800 more) than at lower doses of worms. This appeared to be due to a stronger immune response of the host at higher doses. However, the increase in the rate of rejection was still not as great as the increase in the dose. We postulate that the delay in rejection with increased dose is due to a requirement for a "critical mass" of effectors/worm required to cause rejection. As dose increases, more time is required to reach the level at which worm rejection commences. Deaths due to higher doses of worms also occurred in a strain-specific manner and were temporally biphasic. The intestinal phase of infection produced mortality from 1 to 5 days after infection and the strongest rejection phenotype (NFS) was also the most resistant to intestinal deaths. Deaths occurring after Day 5 were due to the parenterally migrating newborn larvae. The weakest rejection phenotype, that of the B10 congenics, was also the least resistant to intestinal deaths. An experimental formula describing 98% worm rejection time with different doses was derived from the data.(ABSTRACT TRUNCATED AT 400 WORDS)     

Changes in mortality and immunological variables of Litopenaeus vannamei parents and their filial families infected with white spot syndrome under different experimental conditions

In order to find changes in mortality and immunological variables of Litopenaeus vannamei parents and the filial WSSV-resistant and -susceptible families after infection with WSSV under different experimental conditions, the haemolymph total haemocyte count (THC), phenoloxidase (PO), and superoxide dismutase (SOD) activities were measured at days 0, 1, 3, 6, 9, 12 and 15 after challenge and shrimp mortality was also recorded. When shrimps were challenged with 10(-3) (1.29x10(6)copiesmL(-1)), 10(-4) (1.29x10(5)copiesmL(-1)) or 10(-5) (1.29x10(4)copiesmL(-1)) WSSV stock solution (0.1mLshrimp(-1)), the cumulative mortalities (mean+/-S.E.) on day 15 were 100+/-0%, 79.3+/-1.1%, and 21.7+/-2.3%, respectively. Among shrimps challenged with 10(-4) (1.29x10(5)copiesmL(-1)) WSSV dilution (0.1mLshrimp(-1)), the cumulative mortalities (mean+/-S.E.) on day 15 in high-density (100shrimpsm(-3)), middle-density (50shrimpsm(-3)), and low-density (25shrimpm(-3)) groups were 95.5+/-0%, 84.7+/-0%, and 72.3+/-0%, respectively. The immunological variables including THC, PO, and SOD were decreased significantly at the beginning of infection stage, while these immunological variables for survivors reached almost the similar levels to the non-infection control group on day 15 after challenge with 10(-4) (1.29x10(5)copiesmL(-1)) WSSV dilution (0.1mLshrimp(-1)). Cumulative mortality (mean+/-S.E.) on day 15 in 17 filial families (G(2)) ranged from 13.3+/-1.9% to 100+/-0% when shrimps were challenged with 10(-4) (1.29x10(5)copiesmL(-1)) WSSV dilution (0.1mLshrimp(-1)). Although, the PO and SOD activities for shrimps in the WSSV-resistant family were slightly higher than those in the WSSV-susceptible family at the same sampling time after infection, these differences were not significant (p<0.05).     

Reactivating tammar wallaby blastocysts oxidize fatty acids and amino acids.

The tammar wallaby, Macropus eugenii, has a ruminant-like digestive system which may make a significant concentration of amino acids and fatty acids available to the blastocyst via uterine fluids. Fluorescent and radioisotope analyses were performed to determine the rate of glutamine and palmitate use by blastocysts recovered on day 0, 3, 4, 5 and 10 after reactivation induced by removal of pouch young (RPY). Between day 0 and 4 glutamine uptake increased from 15.6 +/- 6.6 to 36.1 +/- 2.7 pmol per embryo h-1 (P < 0.01) and ammonium production increased from 8.2 +/- 4.3 to 26.6 +/- 3.0 pmol per embryo h-1 (P < 0.01). Glutamine oxidation did not increase until day 10 after RPY (P < 0.01), but the percentage of glutamine oxidized increased from 4.5 +/- 3.1% during diapause to 31.2 +/- 12.6% (P < 0.01) by day 5 after RPY and increased further to 51.0 +/- 15.8% (P < 0.01) by day 10 after RPY. Palmitate oxidation also increased from 0.3 +/- 0.1 by day 0 blastocysts to 3.8 +/- 1.7 pmol per embryo h-1 (P < 0.01) by day 4 blastocysts. This increase provides a greater potential for ATP production, possibly to supply increased demand due to the coincident resumption of mitoses. The ATP:ADP ratio within blastocysts had reduced by the time of the first measurement at day 3 (0.5 +/- 0.2 pmol per embryo h-1; P < 0.01) compared with day 0 blastocysts (1.4 +/- 0.3 pmol per embryo h-1). It is likely that metabolism of amino acids and fatty acids contributes to the energy supply during reactivation of tammar wallaby blastocysts after embryonic diapause.     

Anthelmintic effects of bithionol, paromomycin sulphate, flubendazole and mebendazole on mature and immature Hymenolepis nana in mice

The anthelmintic effects of anti-tapeworm drugs, bithionol, paromomycin sulphate, flubendazole and mebendazole on immature and mature Hymenolepis nana in mice were compared. Immature worms were not affected by paromomycin sulphate or flubendazole administered for 12 consecutive days (days one to 12 after infection) at 100 mg/kg/day but 48% and 100% of H. nana were eliminated from mice by bithionol and mebendazole respectively, at the same dosage regimen. Bithionol, paromomycin sulphate, flubendazole and mebendazole given at 100 mg/kg/day for five consecutive days (days 12 to 16 after infection) eliminated 32%, 29%, 36% and 100% of mature worms respectively. 10 and 20 mg of mebendazole/kg/day for five consecutive days (days 12 to 16 after infection) had little effect on mature worms whereas 50 and 100 mg/kg/day for the same period eliminated 99% and 100% of mature worms, respectively. ED50 of mebendazole in the elimination of mature H. nana was 14 or 15 mg/kg/day for five days from the reduction in dry weight or in number of worms recovered respectively. The effects of mebendazole given 2 to 4 days, 8 to 10 days or 13 to 15 days after infection at 100 mg/kg/day were compared. Very low, if any, activity of the drug given 2 to 4 days after infection was seen, whereas the drug given 8 to 10 days or 13 to 15 days after infection eliminated 84% and 86% of H. nana respectively.     

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.     

Establishment, development and fecundity of Taenia crassiceps in the intestine of prednisolone-treated Mongolian gerbils and inbred mice

Worm establishment, development and fecundity of Taenia crassiceps in the intestine of prednisolone (PTBA)-treated, 15- and 4-week-old Mongolian gerbils and 9-week-old inbred mice of 4 strains (AKR/J, BALB/cAn, B10D2/oSn and C57BL/KsJ) were investigated following oral administration of metacestodes. Gerbils were divided into 5 groups of 4 animals each according to the host age and commencement day of PTBA-treatment (day -13, -7 or 0 relative to infection). Worm recovery from the intestine on day 35 postinfection was not affected by host age, but fewer worms were recovered the earlier the commencement of PTBA-treatment. Worm size, determined by wet weight, total length and proglottis number, correlated inversely with worm burden, suggesting they were affected by the crowding effect. Proglottides were released normally in the faeces but were markedly depressed in all groups except for that of young gerbils. Furthermore, egg production and its development in gravid proglottides were markedly depressed in all groups. In PTBA-treated mice of 4 strains, sexually mature but not gravid worms were recovered from all mice of the AKR/J strain on days 20-32 postinfection, but none or few worms from the intestine of the others. PTBA-treatment did not inhibit all protective host defence mechanism(s) in the unnatural or alternative rodent definitive host of T. crassiceps.     

日本血吸虫尾蚴及童虫的生理学比较

本文介绍了经0.05％中性红-氢氧化钾染色确定为活的血吸虫尾蚴和童虫的生理学比较结果。与尾蚴相比,童虫体表对PAS及Alcian蓝阳性反应消失,与抗血清接触不再产生套膜反应;对水不能耐受,在淡水中6小时内已全部死亡,而在生理盐水中仍有92.7％存活;钻腺内含物排空,童虫不能再经皮肤感染,但越过皮肤屏障若被注入皮下或腹腔则分别有45.4—46.5％及56.9—64.9％发育为成虫。     

Brugia pahangi in rats: increasing the number of infective larvae inoculated increases the percentage of microfilaremic rats

One hundred Brugia pahangi infective larvae (L3) caused microfilaremic (mf + ve) infection in 56% of inbred PVG rats. Adult worms were recovered consistently from infected rats but worm recovery was very low, only 1-3% of L3 inoculated survived to adulthood and the worms were dispersed in a wide range of anatomical sites. This suggested that lack of microfilaremia may be due to the low probability of male and female worms meeting in the same site and thus may be numerically and topographically based. When the number of infective larvae inoculated was increased to 500, the percentage of mf + ve infections in rats also increased to 94%, corroborating the hypothesis that lack of mf was not due to an immune response. In a further experiment all infected rats had lost both mf and adult worms by day 420. It has yet to be established whether final rejection of the parasite is due to immunity.     

Brugia pahangi: susceptibility and macroscopic pathology of golden hamster.

Twenty male hamsters were inoculated with 95 to 150 infective larvae of B. pahangi via the subcutaneous route. Worms recovered from 19 hamsters averaged 14% (0–32) from 11 hamsters killed at 105–195 days after infection and 16% (5–19) from 8 hamsters examined at 23–45 days after infection. Approximately one-half of the worms recovered were from the lymphatic vessels of the testes, epididymis, and spermatic cord. A few were found in afferent or efferent vessels of regional lymph nodes. The remaining worms were from the heart and lungs. Low-level microfilaremias were observed in 10 of 12 hamsters held for over 100 days. The average prepatent period was 89 days (65–128). Worms were recovered for up to 3 weeks following inoculation of nine hamsters via the intraperitoneal route with 100–400 infective larvae of B. pahangi.Gross lymphatic pathologic lesions consisted of moderate to marked dilation of lymphatic vessels, enlargement of regional lymph nodes, and numerous lymphthrombi and emboli. Macroscopic changes were most consistent and severe in the lymphatic vessels of the testes, epididymis, and spermatic cord and were noted less frequently in the afferent or efferent vessels of various regional lymph nodes. Areas of reddish discoloration were observed frequently on the serosal surface of the lung in infected hamsters.     

Contamination of red-meat carcasses by Campylobacter fetus subsp. jejuni.

Campylobacter fetus subsp. jejuni was commonly present in the feces of unweaned calves (2 to 3 weeks old) and from two of four groups of sheep. One new season lamb (12 to 16 weeks old) carried the organism, but the bacteria were not isolated from cattle. With unweaned calves, the fractions of animals infected and carcasses contaminated were similar. Contamination of carcasses usually involved low densities of C. fetus subsp. jejuni (ca. 1 to 10/cm2), which were isolated from flank but not rump areas. The organism was recovered less frequently from chilled carcasses and deboned veal. Small numbers of C. fetus subsp. jejuni could be recovered from equipment during the processing of unweaned calves but not after routine cleaning.     

Effects of host age on infection of ICR mice with Echinostoma caproni or E. trivolvis

There was no significant difference in the number of E. caproni recovered 21 days post-infection from 1-, 2-, 4-, 5-, 6-, 7-, 12-, or 21-month-old ICR mice infected with 25 metacercarial cysts. A range of 10.8-17.3 worms was recovered from mice in the age groups. In a similar experiment with E. trivolvis, there was no significant difference in worms recovered in young vs old mice. A range of 0-1.7 worms was recovered from mice in each age group. Contrary to a previous study on E. caproni in NMRI mice, our results indicate that host age does not affect establishment of E. caproni or E. trivolvis in the ICR mouse.     

Role of intestinal goblet cells in the expulsion of Gymnophalloides seoi from mice

Role of intestinal goblet cells (GCs) in the expulsion of Gymnophalloides seoi was studied using 4 strains of mice, ICR, C3H/ HeN, BALB/c, and C57BL/6, after infection with 200 metacercariae isolated from oysters. On day 7 postinfection (PI), significantly higher (P < 0.05) worm recovery rates (WRRs) were observed in ICR (29.5 +/- 12.0%) and C3H/HeN (14.8 +/- 8.2%) than in BALB/c (5.7 +/- 5.3%) and C57BL/6 (0.8 +/- 1.1%) mice. Alteration of the GC mucins was marked in C57BL/6 mice. On day 14 PI, 5.2 +/- 5.2% and 0.6 +/- 0.7% of worms were recovered only from ICR and C3H/HeN mice. When C57BL/6 mice were immunosuppressed with prednisolone, WRR on day 7 PI increased to 11.7 +/- 13.9%, whereas the GC hyperplasia and mucin alteration diminished significantly. The results suggest that expulsion of G. seoi from the intestine is dependent on immune responses of the host, and GCs may be an important effector.