TECHNIQUES APPLIED in JAPAN for the CONTROL of ASCARIS and HOOKWORM INFECTIONS •\A REVIEW•\ Department of Parasitology, Nati

Japan. J. Med. Sci. Biol., 18, 1-17, 1965

TECHNIQUES APPLIED IN JAPAN FOR THE CONTROL OF ASCARIS AND HOOKWORM INFECTIONS •\A REVIEW•\

YOSHITAKA KOMIYA AND AKIO KOBAYASHI

Department of Parasitology, National Institute of Health, Tokyo , Japan

(Received : February 10th 1965)

The quickest way to control and eradicate parasites is to break up their life cycle at its weakest point. In the cases of soil-borne parasites, techniques applied in the break- up of their life cycle include (1) repeated mass treatment of the infected population to eliminate the worms they harbor, (2) handling and treatment of night-soil to render eggs and larvae non-viable, and (3) procedure for the removal or inactivation of eggs on food . Among these, in Japan, the periodic mass treatments are recognized as the •gmain main pillar•h on which prevention of Ascaris and hookworm infections rests.

The following is a brief summary of techniques developed and applied in Japan .

MASS TREATMENT

a) General considerations

In a community, especially in a rural one, mass treatment is used for the purpose

of rapidly reducing the total number of egg-producing worms and consequently of

minimizing the chance of reinfection.

After one mass treatment, the extent of reinfection diminishes and if a second mass

treatment is performed prior to the time when the number of people positive for eggs

reaches the initial pre-treatment level , the total worm burden, as well as the number of

people infected, will decrease gradually. Repeated mass treatment in non-rural communities will also reduce the worm burden , provided that measures are effectively taken to avoid reinfection.

b) Measurement of worm burden

In order to assess the effect of mass treatment it is necessary to measure the total

worm burden in a given community, before and after the treatment . This can be done by egg-counting technique but because of its complexity it is difficult to use it as a

routine method in large-scale surveys. Its weakness lies in the fact that eggs cannot

be recovered in cases of very light hookworm infection since the sample in which eggs are counted •\ usually 10 mg of stool •\ is too small for this.

In Japan, the technique used for routine stool examinations is the direct smear for

Ascaris eggs, and floatation in a saturated salt solution for hookworm eggs . Worm burdens are recorded as follows : + when egg number under the cover glass (18 •~ 18 mm)

is less than 9, ++ when it amounts to 10-99 , and +++ when it is more than 100. The number of people positive for eggs, in relation to the total number of people examined, can be used for indicating roughly the average individual worm burden .

小宮義孝・小林昭夫(国立予防衛生研究所寄生虫部)

1 2 KOMIYA et KOBAYASHI Vol. 18

Autopsies have shown that the average number of Ascaris per positive case was 5.1 when the incidence of infection was 40 %, whereas the majority (about 70 %) of the infected harbored only one worm when the incidence was 10 % or less (Komiya et al., 1963 a). As a result of mass treatment, decline in individual worm burden appears to occur more rapidly than the decline in over-all incidence. A field experiment on hookworm (Komiya et al., 1953) revealed that incidence fell from about 67 % to 13.5 % over a period of three years, whilst the average worm number decreased from the initial 32 to about 1 per positive case. c) Frequency of mass treatment and season in which it should be carried out When incidence is high it is well known that mass treatment for Ascaris infection given once or twice a year is insufficient to reduce the incidence because of the rapidity of reinfection. If the initial incidence is not too high, say 60-70 % or less, biannual treatments appear to be effective. With the assistance of the Gunma Educational Committee, the authors carried out biannual mass treatments for Ascaris infection among children and pupils in fifty schools. Prior to the initial performance of mass treatment 73 % of the children were positive. After one year this incidence was reduced by half (Fig. 1) and continued to be reduced in the same proportion the following years (Gunma Educational Committee, 1954). Table 1 shows the results of repeated mass treatments twice yearly given to children in 32 schools in Shinjuku Ward in Tokyo. Here the initial incidence (11.6 %) decreased to 2.2 % within five years. The effect of repeated mass treatments in controlling hookworm infection is considered to be about the same as that for Ascaris. Ascaris control is even more effective when mass treatment is crried out after the

Fig. 1. Decrease in the incidence of Ascaris infection as a result of biannual mass treatments.

Table 1. Decrease in the incidence of Ascaris infection as a result of biannual mass treatments 1965 CONTROL OF ASCARIS AND HOOKWORM INFECTIONS 3 infection has reached its peak, i. e. in May and October in Japan, when raw vegetables are mostly to be seen in the markets (Fig. 2) (Kobayashi , 1954; Yajima, 1955). In hookworm infection such treatment should be carried out in winter , owing to the low resistance of eggs and larvae to low temperatures (Fig. 3) (Kobayashi et al., 1959). Table 2 shows the results of experimental mass treatments performed in winter over a period of three years (Komiya et al., 1953). Here the initial worm burden (32.1 per the infected) decreased to about 1/30 (1.1 per infected) after two years. d) Performance of mass treatment In order to carry out mass treatment effectively the following is recommended. 1) Stool examinations : The performance of stool examinations of all people in a

Fig. 2. Seasonal variation of Ascaris infection

Fig. 3. The resistance of hookworm eggs to low temperatures Kobayashi et al., 1959)

Table 2. Number of worms expelled after mass treatments (Komiya et al., 1953) 4 KOMIYA et KOBAYASHI Vol. 18 community prior to mass treatment is bound to be difficult, but it is essential that at least 80 % of the population is examined previously for eggs. To achieve this, health education is an absolute necessity. Simple posters, pamphlets and pictures should be utilized. Small groups should be invited to view slides and movies.

2) Technique for the recovery of eggs from stools : For evaluation purposes, stool examinations should be made before and after mass treatment, using a standardized technique.

As mentioned above the two techniques which are recommended in Japan are : the direct smear with a 18 •~ 18 mm cover glass for Ascaris eggs (provided that the same specimen is examined three times) and floatation in a saturated salt solution for hookworm eggs.

The direct smear is sufficient for the recovery of fertilized Ascaris eggs even in mild infection cases. But when incidence is reduced the number of people discharging only unfertilized eggs was found to increase gradually. For instance, when the former become as low as 5 % or less, the latter accounts for 80 % or more of all the positives

(Komiya et al., 1963 a). In such a case those discharging only unfertilized eggs are considered as harboring only one female, and the EPG of such a female is considered to be much lower than the EPG of a female producing fertilized eggs (Komiya et

al., 1963 a).

It seems that the •gthick thick smear technique with cellophane paper cover•h (Kato and

Miura, 1954) enables the recovery of more unfertilized eggs. This technique has been

officially recommended by the Parasite Control Association of Japan for use in a nation-

wide survey of Ascaris and hookworm infections.

3) Selection of anthelmintics : For mass treatment to be successful an anthelmintic

should be selected in advance which must be very effective with few side-effects, inex-

pensive and easily tolerated. With the above in mind the Ministry of Health and Welfare of Japan (1961)

recommended the following anthelmintics for use in mass treatments of ascariasis and

ancylostom iasis. For ascariasis : 1. Santonin 2. A combination of santonin and kainic acid 3. Piperazine preparation 4. A combination of santonin with oil of chenopodium For ancylostomiasis : 1. Tetrachlorethylene 2. 1-bromo-2-naphthol 3. 4-iodothymol 4. Bephenium hydroxynaphthoate Among these santonin with kainic acid and 1-bromo-2-naphthol are particularly recommended for Ascaria and hookworm infections, respectively. Both anthelmintics have been developed in Japan and proved to be highly effective with few side-effects. It is possible that these drugs may not yet be widely used outside the country. Brief inf ormations may therefore be necessary under the separate chapter as to their an thel- mintic activities and toxicities. In some communities, it will be necessary to determine the predominant hookworm species involved prior to drug administration so that a better compound is given. For such a purpose, the cultivation technique using a filter paper (Harada and Mori, 1951) may be conveniently adopted. 1965 CONTROL OF ASCARIS AND HOOKWORM INFECTIONS 5

4) Assessment of mass treatment : In order to assess the results obtained after treatment it is necessary to perform stool examinations. Morishita working on Ascaris

(1953) and Komiya et al. (1954 a) on hookworm have shown that Ascaris is completely expelled within 14 days and hookworms within 4 days. As a safety measure another week should be added to the expulsion period and stool examination should be performed

3-4 weeks after treatment for Ascaris, 2-3 weeks for hookworms, or if convenient 3-4

weeks in both cases.

The anthelmintic action of a given drug against Ascaris or hookworm infection may

possibly be evaluated according to one of the following criteria: (1) the rate of ex-

pulsion of the worms, (2) the egg-reduction rate, and (3) the egg-negative rate. Among these, particularly in the case of large-scale mass treatment, the egg-negative rate i, e.

the percentage of patients who became egg-negative in relation to the number of patients

treated, is generally accepted as practically the best criterion for judging the anthelmintic

effect in this country.

In this respect, however, some difficulties may arise in examination techniques as to

accuracy of detecting ova in stools. There is a greater possibility of missing eggs in

faecal specimens as the total worm burden in a population decreases following mass

treatment. This fact was proved by Komiya et al. (1954 b) both theoretically and

practically. For post-treatment examinations, the authors stressed the need to perform at leastt

three successive examinations using the smear and brine floatation techniques, in order

to prevent or minimize the risk of false negative results following treatment of ascariasis

and ancylostomiasis, respectively. The Harada-Mori's cultivation technique can be utilized

in this respect, in conjunction with the floatation technique but not in cold winter

(Komiya et al., 1960).

e) Anthelmintic effects of the combination of santonin and kainic acid

In Japan, santonin and Digenea simplex Ag. (the dried seaweed) have been preferably

and effectevely used so far against Ascaris infections.

In 1953 Murakami and Takemoto succeeded in separating an effective fraction

(digenic acid) from an extract of Digenea simplex, ultimately designating this principle as •gkainic acid•h. The structural formula of kainic acid is shown below :

Ls-arabo-2-carboxy-3-carboxymethyl-4-isopropenyl-pyrrolidine Thanking to their contribution, coupled with the successful mass production of santonin, Japan has become possible to supply a sufficient amount of santonin and the effect-stabilized Digenea to meet both domestic and foreign demands. Muto and Sasaki (1925) first suggested that santonin and Digenea showed a higher anthelmintic efficacy against ascariasis when used jointly than when used separately. This fact was confirmed later by a number of investigators (Takemoto et al., 1954; Morishita, 1955; Komiya et al., 1954 c, 1955; Kitamoto et al., 1956; Asada et al., 1956; Takemoto, 1957). 6 KOMIYA et KOBAYASHI Vol. 18

Table 3. Results of treatment of ascariasis with santonin, kainic acid and their mixture (Takemoto, 1957)

* K .G.=Kindergarten childrenP.S.=Primary schoolchildren J.H.=Junior high-school pupilsS.H.=Senior high-school students

Takemoto et al. (1954, 1957) tested the ascaricid effects of the mixture of santonin and kainic acid in various dose combinations. Santonin and kainic acid were given separately to control groups. In the control groups, only 20.9 % and 30.8 % of Ascaris- infected children were cured following treatment with 80 mg of santonin and 10 mg of kainic acid, respectively. Among children who were given a mixture of 50 mg santonin and 5 mg kainic acid, however, 68.5 % of 203 persons were cured, and when 50 mg of santonin plus 10 mg of kainic acid was given once, the cure rate was as high as 86.9 % (Table 3). Thus a combined treatment with santonin and kainic acid was found to produce a synergetic action and result in a marked increase in anthelmintic activity. The synergetic action of santonin and kainic acid was then confirmed by Morishita (1955) with similar results. In the light of these facts, 10 parts of santonin to one part of kainic acid is now generally considered as being the best combination. The senior author and his associates (Komiya et al., 1955) tested the efficacy of kainic acid and three mixture preparations of santonin and kainic acid against Ascaris infection, santonin serving as control. The study was composed of three separate tests and the results are given in Table 4.

Table 4. Results of treatment of Ascaris infection with santonin, kainic

*K .a.=Kainic acid 1965 CONTROL OF ASCARIS AND HOOKWORM INFECTIONS 7

In group A, no difference was demonstrated between kainic acid (20 mg) and santonin

(60 mg), the cure rates being 46.5 % and 42.1 %, respectively. In group B, a dose of 100 mg of santonin produced a cure rate of 66.7 %. In contrast, the figures relating to the mixture preparations appeared to be much higher.

The administration of 2 tablets of •gNeo-macnin S•h (50 mg of santonin and 5 mg of

Kainic acid, total) cured 74.2 % of 66 patients. A much higher cure rate -95.4 % - was obtained when 5 tablets of •gDaias•h (100 mg of Santonin and 5 mg of kainic acid, total) were used.

The superiority of the mixture preparation over santonin alone regarding the therapeutic value was also shown in Group C. Here 10 tablets of •gDigeran•h (59 mg of santonin and 1240 mg of Digenea extract, total) produced a 72.1 per cent cure, while the figure for santonin alone was only 47.3 %.

Although some untoward side-reactions were encountered in some of the patients treated with the mixture preparations, the symptoms were mild and transient, and their nature and intensity were almost the same as those produced by treatment with santonin alone. In the group treated with both santonin and the mixture preparations xanthopsia and headache were the main symptoms encountered.

In this paper, an indication will be given of the comparative effects of piperazine and the santonin-kainic acid mixture. Yanagisawa et al. (1957) reported the results of a comparative test on piperazine malate (•gBexin•h) and the santonin-kainic acid mixture

(•gMacnin S•h) in an Ascaris-infected adult population. •gBexin•h was given at a dose of 40 mg/kg of body weight for two days (4.0 g total per person) and •gMacnin S•h at a dose of 50 mg santonin and 5 mg kainic acid for two days. The results are shown in Table 5.

As will be noted, the cure rate for 89 patients treated with •gBexin•h was 86.5 %, while for those treated with •gMacnin S•h the cure rate was 78.5 %. This difference was not significant. Untoward side-reactions such as diarrhoea and loss of appetite were observed in a few patients in each group. However, mild and transient xanthopsia was noted in half of those treated with the mixture preparation.

The treatment of Ascaris infection with a very small dose of the mixture of santonin and kainic acid points to the high efficacy of the preparation. The preparation is highly acceptable to children and is essentially non-toxic. In addition to these advantages the possibility of mass production of the drug at a moderate price facilitates considerably the mass treatment of Ascaris infections.

acid and their mixture preparations (Komiya et al., 1955)

**S=Santonin 8 KOMIYA et KOBA YASHI Vol. 18

Table 5. Comparison of anthelmintic effects of piperazine malate

(•gBexin•h) and santonin-kainic acid mixture (•gMacnin S•h) against ascariasis (Yanagisawa et al., 1957)

* In terms of piperazine hydrate

One tablet of •gBexin•h contains 200mg of piperazine hydrate

In this respect, it may be said that the mixture of santonin and kainic acid is, toge- ther with piperazine compounds, a highly promising anthelmintic against Ascaris infection. f) Anthelmintic effects of 1-bromo-2-naphthol The 1-bromo-2-naphthol is one of the naphthalene derivatives synthetized by Miura and his associates in 1953 in the course of their exploratory studies on anthelmintics active against hookworms. The structural formula of this compound is as follows :

The 1-bromo-2-naphthol, manufactured by Toyama Chemical Industrial Co . Ltd., is now available under the proprietary name of •gWormin•h, in different forms of pure powder, granules and tablets. Of these, the most commonly available one is granules, 1.5 g of which contains 1.0 g of 1-bromo-2-naphthol. In this paper, drug dosages are indicated in terms of the base product, unless otherwise referred to.

After this compound was found, Iwata et al. (1953, 1954) first tried it in the treatment of human hookworm infections and demonstrated it as being highly effective. As an additional advantage, they indicated that 1-bromo-2-naphthol showed a low toxicity even when given without post-treatment purgation.

The comparative anthelmintic effects of 1-bromo-2-naphthol against both species of hookworms, Ancylostoma duodenale and Necator americanus, were determined by several investigators (Matsuzaki, 1957; Yoshida et al., 1960; Hsieh et al., 1960; etc. ).

Matsuzaki (1957) reported a high efficacy of 1-bromo-2-naphthol against both species of hookworm. The drug was given to persons harboring either Ancylostoma or Necator in different dosage schedules according to the age. The results are shown in Table 6.

Of 76 schoolchildren with Ancylostoma, 63 (i. e. 82.9 %) were cured after being treated with 2.0 g of this compound for three consecutive days , while 80.9 % of 141 Necator- infected children were cured with the same dosage. Similar results were obtained in an adult population ; with 4.0 g for three days, the cure rate amounted to 73 .7 % against Ancylostoma and 77.5 % against Necator. However, in the treatment of junior high- school patients who received 3.0 g of the drug for three days , Matsuzaki obtained a 46.7 per cent cure rate in 15 cases with Ancylostoma infection. In contrast, a higher cure rate - 82.4 % - was obtained in 51 Necator-infected cases. 1965 CONTROL OF ASCARIS AND HOOKWORM INFECTIONS 9

Table 6. Difference in resistance against 1-bromo-2-naphthol between two species of hookworms (Matsuzaki, 1957)

* A .d.=Ancylostoma duodenale N.a.=Necator americanus

Yoshida et al. (1960) tested four different anthelmintics : tetrachlorethylene, 4-iodothymol, bephenium hydroxynaphthoate and 1-bromo-2-naphthol against Ancylostoma and Necator infections. From their findings, it was considered that bephenium hydroxynaphthoate would be more effective against Ancylostoma than against Necator, while tetrachlorethylene and 4-iodothymol would produce opposite effects. However, 1-brome- 2-naphthol was found to have an almost equal degree of anthelmintic activity against both species of hookworms. Concerning the comparative activity of 1-bromo-2-naphthol against Ancylostoma and Necator infections, however, somewhat different data were presented by Hsieh et al. (1960) in Formosa. In their study, 1-bromo-2-naphthol and tetrachlorethylene were employed in the treatment of hookworm infections. A dose of 4.0 g of 1-bromo-2- naphthol or 3.0 ml of tetrachlorethylene syrup (adult dose) was administered. From their results, it was found that both drugs were more effective against Necator than Ancylostoma. For the treatment of Necator infection, 1-bromo-2-naphthol resulted in curing as much as 87.5 % of the people, in contrast to the rate of 52.1 % for Ancylos- toma infection. The evidence obtained by Hsieh et al. as to the efficacy of 1-bromo-2-naphthol against Ancylostoma and Necator infections is in full agreement with earlier data (Yamazaki, 1955) obtained by a clinical trial of this compound. In all, it may be safely said that 1-bromo-2-naphthol is effective against both species of hookworms but slightly more so against Necator americanus. A comparative study on the anthelmintic effects of 1-bromo-2-naphthol and bephenium hydroxynaphthoate was carried out by the authors (Komiya et al., 1964) in an area where Necator predominated. In this study, a single dose of 1-bromo-2-naphthol (8.0 g, 6.0 g or 4.0 g) or bephenium hydroxynaphthoate (2.5 g, 1.5 g or 1.0 g as bephenium ion) was given to Necator-infected adults. The results are shown in Table 7. Cure rates obtained were as follows : one dose of 8.0 g of bromo-naphthol - 95.7 % ; 6.0 g of bromo-naphthol - 60.0 % ; 2.5 g of bephenium - 73.5 % ; and 1.5 g of bephenium - 40.8 %. No marked difference was observed between both drugs as regards the occurrence of side-effects. The superiority of 1-bromo-2-naphthol over bephenium hydroxynaphthoate against Necator infections, as evidenced by this study at the recommended doses, was also indicated by Yokogawa et al. (1961) and Juttijudata et al. (1962). 10 KOMIYA et KOBAYASHI Vol. 18

Table 7. Comparison of the effects of 1-bromo-2-naphthol and bephenium hydroxynaphthoate against Necator infection (Komiya et al ., 1954)

Side-effects of 1-bromo-2-naphthol and bephenium hydroxynaphthoate

* Dosages are shown in terms of base products . One gram of 1-bromo-2-naphthol is equivalent to 1.5 g of •gWormin•h granules , and 1.0 g of bephenium ion to 2.0 g of •g Alcopar•h

In the treatment of Ancylostoma infection with these drugs , however, the authors obtained entirely different results from those against Necator as to their comparative efficacies. The data indicated that , as shown in Table 8, 1.5 g and/or 1.0 g of bephenium produced an extremely high cure rate such as more than 90 % . In contrast to this, 6.0 g of bromo-naphthol produced only 68.8 per cent cure. Although the cure rates thus obtained with both drugs possibly be altered by the subjected patients as the worm burden differs from a population to another , their comparative values against each species of hookworms would be qualitatively similar through other cases. Thus, it is our opinion that, as advocated by Yoshida et al . (1960), Fushimi (1961)

Table 8. Comparison of the effects of 1-brome-2-naphthol and bephenium hydroxynaphthoate against Ancylostona infections (Komiya et al., 1964)

* For the dosages refer to the foot note in Table 7 . 1965 CONTROL OF ASCARIS AND HOOKWORM INFECTIONS 11 and Morishita and Fushimi (1962), the better use of the drug would be the administration of 1-bromo-2-naphthol against Necator and bephenium hydroxynaphthoate against Ancylostora. Oshima (1963) performed a study in a Necator area in an attempt to evaluate the anthelmintic efficacy of triclofenol piperazine (Park, Davis Co. ), using 1-bromo-2-naphthol for comparison. Triclof enol piperazine was administered in a single dose of 2.5 g or 1.5 g for two days (3.0 g total), and 1-bromo-2-naphthol was given once in a dose of 6.0 g or 4.0 g. The cure rates obtained were : 68.9 % of 61 patients having received 6.0 g of bromo-naphthol and 49.1 % 4.0 g of this drug. In contrast, when treated with triclofenol piperazine, cures were obtained only in 15 % and 3 %, respectively. The results of another study recently carried out by the present authors (Komiya et al., 1963 b) are given herein. A series of relevant studies with a large variety of dosages and regimens of 1-bromo-2-naphthol were performed for the purpose of establishing the standard therapeutic regimen in mass treatment of Necator infection. The standard regimen should comply with the following requisites : (1) high efficacy at a small dose, (2) few side-effects, and (3) simplicity of medication procedure. A total of 583 adults infected with Necator were examined to determine the proportion of cures and side-reactions after the treatment. No pre- and post-purgations were made, and no restrictions in diet were recommended before the treatment. The results obtained are shown in Tables 9-11. As seen in Table 9, the highest cure rate was seen in Group G (8.0 g, single dose - cure rate 95.7 % ; Group D (10.0 g, b. i. d., at intervals of 1 hr - 80.0 %) came second ; next in order were Group H (6.0 g, single dose - 66.5 %), Group A (10.0 g, b. i. d., at intervals of 2 hrs - 64.3 %), Group E (8.0 g, b. i. d., at intervals of 1 hr - 63.8 %), then Group F, B, I and C. The cure rates in Group A, E and H were almost identical, ranging from 63.8 to 66.5 %. Among groups G, D, H, A and E, in which cures of more than 60 % were obtained, Group G, D, A and E presented a rather high proportion of side-effects - as much as 60 % or more, whereas in Group H side-reactions occurred in only 38 % of the patients (Table 10). Although side-effects occurred in approximately one third of the individuals of Group H, the great majority of the symptoms were transient and mild. The chief complaints in this group were diarrhoea (21.1 %), headache (11.9 %) and abdominal pain

Table 9. Results of treatment of Necator infection with 1-bromo-2-naphthol in various dosage schedules (Komiya et al., 1963b)

* % of cases which showed no eggs in faeces, after three successive brine floatation tests. 12 KOMIYA et KOBAYASHI Vol. 18

Table 10. Side-effects of 1-bromo-2-naphthol in various dosage schedules (Komiya et al., 1963b)

Table 11. Nature and intensity of side-effects in 194 cases treated with a single dose of 6.0 g of 1-bromo-2-naphthol (Komiya et al., 1963b)

(7.2 %) (Table 11). Thus, a single treatment dose of 6.0 g of this compound (9.0 g of •gWormin•h granules) showed the best results as regards the therapeutic value and side-effects. In addition to these advantages, this dosage schedule simplifies the medication procedure and is inexpensive. Taking the above into account, it was concluded that this therapeutic regimen should be recommended as a standard one for the mass treatment of Necator infection, as far as has been tested.

HANDLING AND TREATMENT OF NIGHT-SOIL a) Index of ovicidal effect Among the helminth eggs present in night-soil, those of Ascaris have proved to be the most resistant against chemicals as well as to low temperatures . Therefore the number of dead Ascaris eggs is generally used as an index of the ovicidal effect on helminth eggs in general. b) Storing of night-soil Parasite eggs will gradually degenerate when night-soil stored for a sufficient time undergoes anaerobic decomposition. 1) Storage tanks : The above-mentiomed principle is easily applied . In the majority 1965 CONTROL OF ASCARIS AND HOOKWORM INFECTIONS 13 of farming families in Japan the storage tanks is installed either in the farmyard or in the field ; most Ascaris eggs found in it are killed within a 3-4 week period in summer, when the temperature is usually high, whilst in winter, at low temperature, they are found alive even after 4-5 months. This storage technique of night-soil was particularly recommended in certain rural areas in Japan some 20-30 years ago but is seldom utilized now.

This technique requires, of course, tanks of such a size that the family night-soil can be stored long enough to achieve complete destruction of helminth eggs.

However, one of its defects is that farmers frequently utilize night-soil in a particular season with the result that they consider the storage time recommended is too long.

2) •gKairyo-benjo•h (improved latrine) : The •gKairyo-benjo•h was recommended

about 30 years ago for the purpose of killing helminth eggs in night-soil. Night-soil is stored in it for 5-6 months before use. The tank is divided into three communicating chambers (Fig. 4). All Ascaris eggs are found dead after this time.

If properly maintained, the •gKairyo-benjo•h will contribute towards the control of

Fig. 4. Kairyo benjo (improved latrine for night soil storage)

soil-borne parasites. In practice, however, only 30,000 families (0.5 %) in rural areas

appear to have installed this system due to its high cost, so that in reality it has con-

tributed little towards the control of helminth until now.

c) Compost heaps

In Japan compost is utilized as a fertilizer in rural areas. It is generally composed

of straw, weeds, dead leaves mixed with soil. When human or domestic animal night-

soil is added fermentation takes plase and the temperature may rise to as high as 50-

60•Ž over a period of several days so that most helminth eggs will be killed in due

course. d) Chemical ovicides Several chemicals have been put forward for killing helminth eggs in night-soil, among which are carbon disulfide, mustard oil and sodium nitrite. 1) Carbon disulfide (CS2) : The ovicidal effect of carbon disulfide was initially 14 KOMIYA et KOBA YASHI Vol. 18

reported by Matsumura et al . (1953), who recommended it on account of its inexpen- siveness. When mixed in night-soil in a proportion of about 0 .5 % it kills all Ascaris eggs after seven days at room temperature . Subsequently, the ovicidal effect of carbon disulfide was found to decrease considerably at low temperature (a proportion of

about 5 % of night-soil being necessary for complete killing at 5•Ž) . The authors hesitate to recommend it to the public because of the possible harmful effects of health

and inflammability of the fumes.

2) Mustard oil: The ovicidal effect of mustard oil , first reported by Kunii & Ikeuchi (1953), is excellent in an acid medium but markedly less so in an alkaline

medium as such is usually the case in night-soil . Because of its harmful effects on the human skin and because it is expensive , its use is not recommended. 3) Sodium nitrite (NaNO2) : The ovicidal effect of sodium nitrite on Ascaris eggs

in an acid medium was initially reported by Nagano et al . (1952). It is inexpensive, but its utilization in night-soil meets with difficulty due to the alkaline nature of the

latter.

Recently Kozai (1960) succeeded in keeping the pH of night-soil below 7 .0 by mixing it with 2-5 per cent calcium superphosphate. Not only was no harm done to the crops

but the effect of night-soil as a fertilizer was increased by adding this compound , itself a fertilizer.

By adding sodium nitrite to night-soil previously treated with calcium superphosphate , all Ascaris eggs in night-soil will be killed in three days . The concentration of sodium

nitrite to be added is 0.025-0 .125 % (w/v) of night soil at a temperature range of 26- 29•Ž and 0.1 % at 3-5•Ž.

The authors consider that this technique is very promising . 4) Thiabendazole : Recently Kutsumi (1963) reported that thiabendazole had a high

ovicidal effect in alkaline night-soil . On mixing it with alkaline night-soil (25 p. p. m. of the latter), it killed all the Ascaris eggs at a temperature of 15•Ž after 7 days'

exposure.

OTHER PREVENTIVE MEASURES

In this connexion the following measures are worthy of attention ; a) Synthetic detergents Jn Japan several synthetic detergents specially prepared for food-stuff, most of which are of ABS origin, have been manufactured for the public purposes . All were submitted to the Department of Parasitology , the National Institute of Health, for examination with respect to their effects on Ascaris eggs. When correctly applied (concentration 0.5 %, immersion time 5-10 minutes and washed in the solution after immersion) all types of detergent removed almost all of Ascaris eggs from vegetables (Kobayashi & Kumada, 1957). b) Irradiation The utilization of gamma-rays for the sterilization of Ascaris eggs on vegetables has been given much attention in Japan in recent years. Basic investigations were made on the effects of Co60 irradiation on Ascaris eggs at various developmental stages (Asami et al., 1955; Saito, 1957; Kobayashi et al., 1958). A dose of 153,000 r completely inhibited the embryogenesis of unicellular eggs . To achieve complete inhibition of infectivity or motility of embryonated eggs, however, much 1965 CONTROL OF ASCARIS AND HOOKWORM INFECTIONS 15

higher doses -1, 000, 000 r or more were required. The results appear to offer the possibility of applying ionizing irradiation to vegetables or other food for sterilization purposes.

REFERENCES

ASADA, J., AOKAGE, K., KAJI, F., OCHI, G. & ð•TAGAKI, H. (1956): Treatment of ascariasis

with kainic acid, the crystallized effective principle of Digenea extract, •gTð¯kyð¯-Iji-Shinshi•h , 73, 311-312 (In Japanese)

ASAMI, K., KOBAYASHI, A. & SAITð•, S. (1955): Observations on the ovicidal effects of irra-

diation with cobalt-60 on the development of Ascaris lumbricoides ova. 1. Jap. J. Parasit.,

4, 331-336 (In Japanese with English summary) FUSHIMI, J. (1961): A new therapeutic of hookworm disease. Anthelmintic efficacy of bephe-

nium hydroxynaphthoate on human hookworm infection (Paper presented to the symposium in the local Meeting Jap. Soc. Parasit.)

Gunma Educational Committee, Gunma Health Committee for School (1954): Report on the survey on parasites among schoolchildren. 1. •gGakko-Hoken-Kenkyu•h 11, 1-5 (In Japanese)

HARADA, Y. & MORI, O. (1951): A simple method for the cultivation of hookworm larvae , •g Igaku-to-Seibutsugaku•h, 20, 65-67 (In Japanese) HSIEH, H., CHOW, L. & SHIN, C. (1960): Mass treatment of hookworm infections with tetra-

chlorethylene and 1-bromo-2-naphthol in two rural villages in Southern Taiwan, J. Formosan Med. Ass., 59, 53-57.

IWATA, S., NAKAMURA, I., SHIMADA, H., MIURA, G. & OGATA, K. (1953): 1-bromo-2-

naphthol, a new anthelmintic active against human hookworm, and its therapeutic value,

•g Shinyaku-to-Rinshð¯•h, 2, 354-360 (In Japanese) IWATA, S., NAKAMURA, I. & MIURA, G. (1954): Mass treatment of hookworm infection with

1-bromo-2-naphthol,•gShinyaku-to-Rinshð¯•h, 3, 261-271 (In Japanese)

JUTTIJUDATA, P., GUPTAVANIJ, P. & HARINASUTA, C. (1962): A comparison of the efficacy of anthelmintic drugs in the treatment of hookworm infection, Japan. J. med. Sci. Biol., 15,

61-65.

KATO, T. & MIURA, M. (1954): On the comparison of some stool examination methods, Jap. J. Parasit., 3, 35 (In Japanese)

KITAMOTO, O., TAKAHARA, T., TAKAYAMA, H., ABE, S., OKADA, K., TANAKA, S., TA- NAKA, T. & ISHIZAKI, T. (1956): Clinical studies on the anthelmintic properties against

ascariasis. 1. The effect of Kaininsan, Santonin and their combination, Jap. J. Parasit., 5, 216 (In Japanese)

KOBAYASHI, A. (1954): Studies on the modes of natural Ascaris infection in Gunma Perfec- ture. 1. On the seasonal variation of Ascaris infection among farmers , •gKitakantð¯ Med. J.•h, 3, 194-199 (In Japanese)

KOBAYASHI, A. & KUMADA, M. (1957): The effect of several syndets on the elimination of Ascaris ova attached to vegetables, Jap. J. Parasit., 6, 491-498 (In Japanese with English

summary)

KOBAYASHI, A., KUMADA, M. & KOMIYA, Y. (1958): Observations on the ovicidal effects of irradiation with cobalt-60 on Ascaris ova. 3. Effects on Ascaris suilla ova in embryonated

stages, Jap. J. Parasit., 7, 39-47 (In Japanese with English summary) KOBAYASHI, A., SUZUKI, N., YASUDA, I., KUMADA, M., ISHII, T., OGAWA, H. & KOMIYA,

Y. (1959): On the resistance of hookworm eggs to lower temperatures, Jap. J. Parasit ., 8, 637-641 (In Japanese with English summary)

KOMIYA, Y., AIZAKI, T., ð•TAKE, S. & TSUKAGOSHI, K. (1953): The field model experiment of hookworm control by repeated mass treatment in winter, Jap. J. Parasit., 2, 157-163 (In

Japanese) 16 KOMIYA et KOBAYASHI Vol. 18

KOMIYA, Y., SATð•, S. , KOJIMA, K., YOKOGAWA, M. , SANO, M., KIHATA, M. & NAGAI, R. (1954a): The general aspect of discharging hookworm and their ova after the administ-

ration of various anthelmintics, Jap. J. Parasit., 2, 221-227; 3, 153-156 (In Japanese) KOMIYA, Y. & SATð•, S. (1954b): The recovery rate of Ascaris and hookworm ova by the

direct faecal smear method, and the rate of false negative cases for ova, in relation to the

assay of the effectiveness of anthelmintics. 1, 2, Jap. J. Parasit., 3, 216-219; 3, 260-264

(In Japanese)

KOMIYA, Y., SATð•, S., YOKOGAWA, M. , AIZAKI, T. & •¬TAKE, S. (1954c): Effects and side- effects in the mass treatment of Ascaris infection with various anthelmintics,•gSaishin-Igaku•h,

9, 1629-1638 (In Japanese) KOMIYA, Y., KONDð•, S., •¬TAKE, S. & TSUKAGOSHI, K. (1955): Anthelmintic effects of kainic

acid and Santonin-kainic acid mixture preparations against Ascaris infection •gSð¯gð¯-Igaku•h,

12, 163-166 (In Japanese) KOMIYA, Y., KOBAYASHI, A., SUGIYAMA, T. & KUTSUMI, H. (1960): Comparative studies on

the technics for stool examination for hookworm ova during the cold climate, particularly on

the recovering rate of hookworm ova with brine floatation technic and on culturing one with filter paper, Jap. J Parasit., 9, 480-485 (In Japanese with English summary) KOMIYA, Y., KOZAI, I. & SUZUKI, R. (1962): The increase of the ratio of those expelling only

unfertilized eggs of Ascaris as the rate of the positive for Ascaris eggs among the social

groups diminishes, and the utilization of this fact in the eradication programme of ascariasis in Japan, Jap. J. Parasit., 11, 45-52 (In Japanese with English summary) KOMIYA, Y., SUZUKI, N, & SUZUKI, R. (1963a): The E. P. G. of a female Ascaris producing

only unfertilized eggs, Jap. J. Para sit., 12, 312 (In Japanese) KOMIYA, Y., KOBAYASHI, A., KUMADA, M., KUTSUMI, H. & SUGIYAMA, T. (1963b): Trial

of establishing the standard therapeutic regime with 1-bromo-2-naphthol (Wormin) in the mass treatment of hookworm infection and comparison of anthelmintic efficacies of two different

forms of the compound, granule and capsule, Jap. J. Parasit., 12, 415-425 (In Japanese with English summary)

KOMIYA, Y., KOBAYASHI, A., KUMADA, M., KUTSUMI, H., IIJIMA, T., ITO, Y. & SUGIYAMA, T. (1964): Comparative tests on the anthelmintic effects of 1-bromo-2-naphthol (Wormin) and

bephenium hydroxynaphthoate (Alcopar) against Necator americanus and Ancylostoma duode-

nale, Jap. J. Parasite., 13, 412-420 (In Japanese with English summary) KOZAI, I. (1960): Re-evaluation of sodium nitrite as the ovicide used in night-soil, 1, 2, 3, Jap.

J. Parasit., 9, 202-210; 9: 519-528; 9, 529-540 (In Japanese with English summary) KUNII, Y. & IKEUCHI, M. (1953): Study on parasite control in villages. The treatment of

Ascaris eggs in night-soil with mustard oil, Bull. Nat. Inst. Agr. Sci., H-6, 55-61 (In Japa- nese)

KUTSUMI, H. (1963): Ovocidal effect of Thiabendazole on Ascaris eggs. 2. Jap. J. Parasit., 12, 303 (In Japanese)

MATSUMURA, T., •¬SAWA, M., ODA, T., NAKAZAWA, S. & YUGAMI, S. (1953): The possible control of Ascaris infection by the treatment of night-soil with chemicals, •gKð¯shð¸-Eisei•h,

13, 40-44 (In Japanese)

MATSUZAKI, Y. (1956): 1-bromo-2-naphthol, an anthelmintic for hookworm,•gShinyaku-to- Rinshð¯•h, 6, 112-114 (In Japanese)

MIURA, K., IKEDA, M., OHASHI, T. & YASUDA, S. (1953a): Studies on anthelmintics. 4. Anthelmintic efficacy of halogenated naphthalene compounds on worms in toads, •gJð¸zen-

Igaku-Zasshi•h, 55, 577-582 (In Japanese) MIURA, K., OHASHI, T. & YASUDA, S. (1953b): Studies on anthelmintics. 5. Anthelmintic

Power of bromonaphthols and their related compounds on toad worms,•gKanazawa-Daigaku- Yakukenhð¯•h, 3, 34-41 (In Japanese)

MIURA, K. (1954): Studies on chemotherapy against hookworm disease. On the effect of 1-

bromo-2-naphthol, Jap. J. med. Sci. Biol., 7, 265-273. CONTROL OF ASCARIS AND HOOKWORM INFECTIONS 1965 17

MORISHITA, K. (1953): Biological studies on Ascaris treatment , Jap. J. Parasit., 2, 13-16 (In Japanese) MORISHITA, K. (1955): A general view of anthelmintics for ascariasis ,•gShinryð¯•h, 8, 585-594 (In Japanese) MORISHITA, K. & FUSHIMI, J. (1952): Anthelmintic effects of bephenium hydroxynaphthoate against hookworm. 3. Efficacies of bephenium against Ancylostoina and Necator , •gShinryð¯•h, 1 5, 1452-1469 (In Japanese)

MURAKAMI, S. TAKEMOTO, T . & SHIMIZU, Z. (1953a): Studies on the effective principles of Digenea simplex. 1. Separation of the effective fraction by liquid chromatography , J. Pharm. S oc. Japan, 73, 1026-1028 (In Japanese)

MURAKAMI, S., TAKEMOTO, T., SHIMIZU, Z . & DAIGO, K. (1953b): Studies on the effective principles of Digenea simplex. 3. Treatment of ascariasis by digenic acid , J. Pharm. ,Soc. J apan, 73, 1055-1057 (In Japanese)

MUTð•, S. & SASAKI, S. (1925): Clinical trial of an anthelmintic (Digeraxine) for the therapy of ascariasis, •gAichi-Igakukai-Zasshi•h , 32, 153-171 (In Japanese) •¬ SHIMA, T. (1963): Unpublished data (personal communication)

SAIT•¬, S. (1957): Observations on the effects of irradiation with cobalt -60 on the development of Ascaris luinbricoides ova. 2. Differences of sensitivity in various developmental stages , J ap. J. Parasit., 6, 175-181 (In Japanese with English summary)

TAKEMOTO, T., SHIMIZU, Z., DAIGO , K. & SAT, T. (1954): Results of treatment of ascariasi s with a mixture of digenic acid and santonin , J. Pharm. Soc. Japan, 74, 107-109 (In Japa- nese) TAKEMOTO, T. (1957): Studies on the effective principles of Digenea simplex , Pamphlet 1-10 (In Japanese) YAJIMA, T. (1955) : Studies on the natural and experimental infections of Ascaris lumbricoid es in man. 1. Studies on the natural infection of Ascaris in man , Jap. J. Parasit., 4, 23-29 ( In Japanese with English summary)

YAMAZAKI, T. (1955) : Treatment of hookworm disease with 1-bromo-2-naphthol ,•gShinyaku- to-Rinshð¯•h, 4, 482-487 (In Japanese)

YANAGISAWA, R., YAJIMA, F. & HARUTA, T. (1957): Anthelmintic effect of piperazine malate

(Bexin) against ascariasis,•gRinsho-Shokakibyogaku•h , 5, 41-42 (In Japanese) YOKOGAWA, M., •¬KURA, T., TSUJI, M., INASAKA , N., NISHI, S. & KIHATA, M. (1961): Mass treatment of hookworm infection with bepheniumn hydroxynaphthoate (Alcopar) , •gIgaku-T sushin•h, 16, 649-653 (In Japanese) YOSHIDA, Y., NAKANISHI, Y., SHIMATANI , T., MATSUO, K. (1960): Comparative studies on the anthelmintic effect of bephenium hydroxynaphthoate , tetrachlorethylene, 1-bromo-2-naphthol and 4-iodothymol against human hookwoom , Jap. J. Parasit., 9, 620-628 (In Japanese)