Formalin-ether Sedimentation Technique Research Articles

Epidemiological Survey on the Infection of Intestinal Flukes in Residents of Muan-gun, Jeollanam-do, the Republic of Korea

Infection status of intestinal flukes was investigated in residents of Muan-gun, Jeollanam-do, the Republic of Korea. Total 1,257 fecal samples of residents were examined by formalin-ether sedimentation technique and Kato-Katz thick smear method. Helminth eggs were detected from 95 (7.6%) residents, and eggs of heterophyid flukes and Clonorchis sinensis were found from 62 (4.9%) and 40 (3.2%) cases, respectively. The larger heterophyid eggs, somewhat dark-brown in color and 37.7 x 21.5 microm in average size, and found in 32 (2.6%) out of 62 egg positive cases of heterophyid flukes. To confirm the adult flukes, we performed worm recovery from 12 cases after praziquantel treatment and purgation with MgSO(4). A total of 1,281 adult flukes, assigned to 7 species, were recovered from 9 cooperative cases. Heterophyes nocens (total 981 specimens) was collected from 9 cases, Stictodora fuscata (80) from 7, Gymnophalloides seoi (75) from 5, Pygidiopsis summa (140) from 3, Stellantchasmus falcatus (3) from 2, and Stictodora lari and Acanthotrema felis (each 1 worm) from 1 case each. The intrauterine eggs of S. fuscata collected from the recovered worm were identical with the larger heterophyid eggs detected in the stool examination. By the present study, it was confirmed that A. felis is a new intestinal fluke infecting humans, and residents in Muan-gun, Jeollanam-do are infected with variable species of intestinal trematodes.

Prevalence of intestinal parasites among primary school children in Makurdi, Benue State- Nigeria

In order to identify the prevalence of intestinal parasitic infections and to determine the impact of some factors such as age, sex and complaints related to these infections, this cross sectional study was carried out on 1000 stool specimen of primary school children in Makurdi, Benue State-Nigeria. Using direct smear and formol ether sedimentation techniques to process the faeces, 585(58.5%) of the samples were found positive for various intestinal parasites with Hookworm species accounting for 34.2%; Ascaris lumbricoides 16.1%; Trichuris trichiura 15.9%; Enterobius vermicularis 6.2%; Strongyloides stercolaris 1.7%; Taenia spp 3.9%; Hymenolepis nana 1.8%; Schistosoma mansoni 0.6%; Entamoeba histolytica 2.2%; Entamoeba coli 1.5%; Giardia lamblia 1.3%. Cases of polyparasitism were also detected but none of the pupils had more than three parasite species. Sex and age did not affect the pattern of infection (P>0.05) since the parasites were similarly found in both sexes and all age groups though with observable varying degree. Results also showed that the average school pupil in Makurdi carry heavy intestinal parasites burden. It is concluded that sanitary measures and deworming program should be conducted in primary schools to decrease the rate of intestinal parasitic infections.

Parasitoses intestinais em um centro de educação infantil público do município de Blumenau,SC, Brasil, com ênfase em Cryptosporidium spp e outros protozoários

The purpose of this study was to establish the prevalence of Cryptosporidium spp and others parasites among infants of 0 to 6 years of age, from a public day-care center in Blumenau, SC, Brazil, between March and May of 2006, through parasitological examinations. The infants’ parents answered a domiciliary inquiry about sanitary conditions and 53 fecal processed samples were evaluated by the formalin-ether sedimentation technique; spontaneous sedimentation, a modified Ziehl-Neelsen, and the ELISA method (TechLab?) for detection of Cryptosporidium parvum. Of the analyzed children it was verified a prevalence of intestinal parasitism of 39,6%, and the parasites found were: Giardia duodenalis (18,9%), Entamoeba histolytica/ Entamoeba dispar/Entamoeba hartmanni (15,1%), Cryptosporidium spp (7,6%), Entamoeba coli (3,8%) Endolimax nana (3,8%) and Cyclospora cayetanensis (1,9%). The methodology, modified Ziehl-Neelsen, used for sample analysis and coloring the structures, was effective on coloring the acid-fast stain structures. The positive cases received specific treatment and the children took part of educational activities. Our data showed that the high rate of parasitism found reveals the necessity to adopt sanitary educational measures towards the people from that area.

A field evaluation of formalin–gasoline technique in the concentration of stool for detection of intestinal parasites

A field evaluation of the formalin-gasoline procedure to detect parasite ova, cysts, or larvae from 470 fecal specimens (comprising both fresh and formalin-preserved stool samples) was compared with that of the formalin-ether sedimentation technique. Parallel concentrates with diethyl ether and gasoline were prepared for each specimen, and the species and appearance of recovered parasite species were determined. Of 470 total specimens, 206 (43.83%) were found to be positive for parasites in one or both concentration techniques. Gasoline was comparable to diethyl ether in the recovery of parasite eggs, cysts, and larvae, so that the formalin-gasoline and the formalin-diethyl ether sedimentation techniques detected 165 and 156 positive of total specimens, respectively. In this study, gasoline proved to be as good as diethyl ether in concentrating parasite eggs and cysts, as well as in maintaining characteristic morphology. However, gasoline was considerably superior to diethyl ether in detecting larvae of Strongyloides stercoralis. Parallel examination of total stool samples by the routine formalin-ether (original) and by the formalin-gasoline techniques resulted in identical distribution of positive slides and morphology of recovered parasite species. However, the easy availability of gasoline (wherever gas stations are present) and its low cost in comparison to ether makes gasoline superior to ether for use in concentration of stools by the sedimentation method in laboratories, including laboratories with limited material resources and also laboratories present in small cities and rural health centers.

Evaluation of techniques for detection of small trematode eggs in faeces of domestic animals

This study was conducted to evaluate techniques for detection of small trematode eggs in faeces of dogs, cats and pigs. Faecal samples from dogs ( n = 80), cats ( n = 35) and pigs ( n = 114) were examined by Kato-Katz technique (KK), formalin–ether sedimentation technique (FE) and a method of combining: filtration, sedimentation and centrifugation, developed at DBL – Centre for Health Research and Development (former Danish Bilharziasis Laboratory) (DBL). Necropsy was performed on 38 dogs, 25 cats and 16 pigs and was considered as a gold standard method for evaluation of the techniques. The results showed 100% specificity for the three techniques. Lower sensitivity was seen for the KK-technique in dog samples in comparison to that for DBL- and FE-technique. The sensitivity of the three techniques was similar in cats and pigs. Based on these findings and practical issues, DBL-technique was chosen as most suitable because the eggs were easily detected and quantified. No toxic chemicals or special equipment were required in comparison with FE-technique that needs ether solution and thus fume cupboards which are often unavailable in local veterinary centres in Vietnam and other developing countries.

Decrease of Metagonimus yokogawai Endemicity along the Tamjin River Basin

The Tamjin River which flows from Jangheung-gun via Gangjin-gun to the South Sea was reported to be a highly endemic area of Metagonimus yokogawai infection in 1977 and 1985. However, there were no recent studies demonstrating how much change occurred in the endemicity, in terms of prevalence and worm burden, of metagonimiasis in this river basin. Thus, a small-scale epidemiological survey was carried out on some residents along the Tamjin River basin in order to determine the current status of M. yokogawai infection. A total of 48 fecal samples were collected and examined by the Kato-Katz thick smear and formalin-ether sedimentation techniques. The egg positive rate of all helminths was 50.0%, and that of M. yokogawai was 37.5%, followed by C. sinensis 22.9% and G. seoi 4.2%. To obtain the adult flukes of M. yokogawai, 6 egg positive cases were treated with praziquantel 10 mg/kg in a single dose and purged with magnesium sulfate. A total of 5,225 adult flukes (average 871 specimens per person) of M. yokogawai were collected from their diarrheic stools. Compared with the data reported in 1977 and 1985, the individual worm burdens appeared to have decreased remarkably, although the prevalence did not decrease at all. It is suggested that the endemicity of M. yokogawai infection along the Tamjin River has been reduced. To confirm this suggestion, the status of infection in snail and fish intermediate hosts should be investigated.

Prevalence of Clonorchiasis in Southern Endemic Areas of Korea in 2006

This study was performed to investigate prevalence of clonorchiasis among the inhabitants living in villages along the 4 major rivers, Nakdong-gang (=river), Seomjin-gang, Youngsan-gang, and Guem-gang in southern Korea. From January to December 2006, a total of 24,075 stool samples (1 sample per an inhabitant) were collected in 23 localities and examined by the formalin-ether sedimentation technique. Of the inhabitants examined, 3,441 (14.3%) were found to harbor various types of intestinal parasite eggs, cysts or larvae. Numbers of infected people were 2,661 (11.1%) for Clonorchis sinensis, 431 (1.8%) for heterophyids, 226 (0.9%) for Entamoeba spp., 57 (0.2%) for Giardia lamblia, 30 (0.1%) for Trichuris trichiura, and 18 (0.07%) for echinostomes. Prevalence rates of clonorchiasis according to the river basin were 17.1% in Nakdong-gang, 11.2% in Seomjin-gang, 5.5% in Youngsan-gang and 4.6% in Guem-gang. Of the 2,661 C. sinensis egg-positive cases, 57.7% was male. The present findings suggest that clonorchiasis is still highly prevalent among inhabitants in the riverside areas of southern Korea, and it is necessary to implement a systematic control program in the endemic areas.

Collection of Clonorchis sinensis adult worms from infected humans after praziquantel treatment

A cohort was established for evaluation of cancer risk factors in Sancheong-gun, Gyeongsangnam-do, Korea. As one of the cohort studies, stools of 947 residents (403 males and 544 females, age range: 29-86 years) were screened for Clonorchis sinensis eggs using both Kato-Katz method and formalin-ether sedimentation technique. The overall egg positive rate of C. sinensis was 37.7% and individual EPG (eggs per gram of feces) counts ranged from 24 to 28,800. Eight egg positive residents voluntarily joined a process of collection of the passed worms after praziquantel treatment. A total of 158 worms were recovered from 5 of the 8 treated persons, ranged from 3 to 108 in each individual. The worms were 15-20 mm x 2-3 mm in size, and showed brown-pigmented, red, or white body colors. This is the first collection record of C. sinensis adult worms from humans through anthelmintic treatment and purgation. The adult worms of C. sinensis may be paralyzed by praziquantel and then discharged passively through bile flow in the bile duct and by peristaltic movement of the bowel.

A new endemic focus of Heterophyes nocens and other heterophyid infections in a coastal area of Gangjin-gun, Jeollanam-do

To know the prevalence of heterophyid trematodes among inhabitants of a southern coastal village, i.e., Sacho-ri, Gangjin-gun, Jeollanam-do (Province), 82 stool samples were examined on helminth eggs and protozoan cysts using Kato-Katz and formalin-ether sedimentation techniques. Total 33 people (40.2%) were positive for trematodes (Heterophyes nocens 15 people, Pygidiopsis summa 3, Metagonimus sp.; 4, Clonorchis sinensis 7, Gymnophalloides seoi 6) and/or protozoa (Entamoeba coli 3). Among intestinal trematode egg positive cases, 17 were treated with praziquantel and their whole diarrheic stools were collected after purgation. Adult flukes of H. nocens (number of specimens = 1,294), P. summa (386), Stellantchasmus falcatus (5), Stictodora lari (4), and Heterophyopsis continua (1) were collected using a stereomicroscope. To know the source of human H. nocens infections in this village, metacercarial infections in mullets (10) were examined and most H. nocens metacercariae (101/105, 96.2%) were found in the trunk portion. From above results, the surveyed coastal village has been newly known as an endemic focus of human H. nocens infection and consuming raw mullets was the presumable source of human heterophyid infections.

A new endemic focus of Gymnophalloides seoi infection on Aphae Island, Shinan-gun, Jeollanam-do

A new endemic focus of Gymnophalloides seoi infection has been discovered on Aphae Island (Shinangun, Jeollanam-do), Republic of Korea. This area, which is referred to as Bokyong-ri, is a small seashore village located in the northern portion of the island. Fecal samples were collected from a total of 57 residents and examined by the Kato-Katz and formalin-ether sedimentation techniques. Helminth eggs were detected in 37 samples (64.9%); 33 samples were positive for G. seoi eggs (57.9%), 4 for Pygidiopsis summa (7.0%), 13 for other heterophyids (22.8%), 1 for Clonorchis sinensis (1.7%), and 1 for Trichuris trichiura (1.7%). Women (70.4%) revealed higher rates of G. seoi infection than did men (46.7%), and individuals older than 50 years of age also evidenced higher rates of G. seoi infection than in other age groups (P < 0.05). In worm collection from 13 G. seoi egg positive cases, G. seoi (total 86,558 specimens), Heterophyes nocens (278), Stictodora sp. (10), Heterophyopsis continua (3), P. summa (3), and unidentified flukes (42) were collected. Oysters, the source of G. seoi infection, were collected from an area proximal to the village and 50 were examined for metacercariae; 47 (94%) were found infected and the observed metacercarial density was 9.5 +/- 8.9 per oyster. The results of this study indicate that Bokyong-ri is a new endemic area of G. seoi infection, with high rates of infection in humans and oysters.

Targeting newly enrolled low-age school children for the control of the intestinal helminth infection in rural Nepal

This study aims to identify an effective intervention group for the control of the intestinal helminth infection among school-age children in rural Nepal. We examined 1677 stool samples of school-age children from 25 schools and 1014 samples from 25 communities in rural Nepal. We used formalin-ether sedimentation technique for the microscopic examination of the stool samples and identified three major intestinal helminths: Ascariasis lumbricoides, hookworm and Trichuris trichiura. Our results revealed that the newly enrolled under-6-year-old children at the schools were more widely infected with at least one of the three major helminths than those in the communities (odds ratio [OR] 2.29). We detected a significant difference in the prevalence of A. lumbricoides (OR 1.70), hookworm (OR 2.57) and T.trichiura (OR 3.23) between under-6-year-old children at the schools and those at the communities. The study results suggest that an appropriate deworming programme is needed for the newly enrolled under-6-year-old children in the primary schools in Nepal.

Comparison of three microscopic techniques for diagnosis ofCyclospora cayetanensis

Cyclospora cayetanensis oocysts in the feces of humans from Kathmandu, Nepal were identified on the basis of their size and other morphological characteristics. We compared the detection of C. cayetanensis oocysts in the feces using three microscopic techniques such as formalin-ether sedimentation, sucrose centrifugal floatation, and direct smear. Standard procedures were used for the formalin-ether sedimentation and the sucrose centrifugal floatation techniques using 0.5 g of feces, however, the direct smear technique was performed using 10 microl of fecal suspension (0.005 g of feces) and observed under the fluorescent microscope. Of the 403 samples examined, 21 samples were positive for oocysts by all three techniques. Therefore, in these 21 samples, the number of oocysts recovered by the three techniques were compared. The highest number of oocyst was obtained by the sucrose centrifugal floatation technique. In contrast, the formalin-ether sedimentation technique was found to be the least reliable concentration technique for the detection of Cyclospora in human feces. Surprisingly, the direct smear technique was found to be an effective and rapid technique for diagnosis of C. cayetanensis making it a technique of choice for routine epidemiological investigation of the prevalence of this infection in human populations.

Comparison of three microscopic techniques for diagnosis of Cyclospora cayetanensis

Cyclospora cayetanensis oocysts in the feces of humans from Kathmandu, Nepal were identified on the basis of their size and other morphological characteristics. We compared the detection of C. cayetanensis oocysts in the feces using three microscopic techniques such as formalin–ether sedimentation, sucrose centrifugal floatation, and direct smear. Standard procedures were used for the formalin–ether sedimentation and the sucrose centrifugal floatation techniques using 0.5 g of feces, however, the direct smear technique was performed using 10 μl of fecal suspension (0.005 g of feces) and observed under the fluorescent microscope. Of the 403 samples examined, 21 samples were positive for oocysts by all three techniques. Therefore, in these 21 samples, the number of oocysts recovered by the three techniques were compared. The highest number of oocyst was obtained by the sucrose centrifugal floatation technique. In contrast, the formalin–ether sedimentation technique was found to be the least reliable concentration technique for the detection of Cyclospora in human feces. Surprisingly, the direct smear technique was found to be an effective and rapid technique for diagnosis of C. cayetanensis making it a technique of choice for routine epidemiological investigation of the prevalence of this infection in human populations.

Status of intestinal parasite infections among children in Bat Dambang, Cambodia

A survey was conducted to determine the extent of intestinal parasite infection in Bat Dambang, Cambodia in March 2004. A total of 623 fecal specimens was collected from kindergarten and schoolchildren and examined using the formalin-ether sedimentation technique. The overall infection rate of intestinal parasites was 25.7% (boys, 26.2%; girls, 25.1%), and the infection rates of intestinal helminthes by species were as follows: Echinostoma sp. 4.8%, hookworm 3.4%, Hymenolepis nana 1.3%, and Rhabditis sp. 1.3%. The infection rates of intestinal protozoa were; Entamoeba coli 4.8%, Giardia lamblia 2.9%, Iodamoeba butschlii 1.4%, Entamoeba polecki 1.1%, and Entamoeba histolytica 0.8%. There were no egg positive cases of Ascaris lumbricoides or Trichuris trichiura. All children infected were treated with albendazole, praziquantel, or metronidazole according to parasite species. The results showed that intestinal parasites are highly endemic in Bat Dambang, Cambodia.

Status of intestinal parasites infection among primary school children in Kampongcham, Cambodia.

A survey was made to find the extent of intestinal parasite infection in Kampongcham, Cambodia in February 2002. A total of 251 fecal specimens were collected from Tonlebat primary school children and examined by formalin-ether sedimentation technique. The overall infection rate of intestinal parasite was 54.2% (males, 57.3%; females, 50.8%). The infection rate of intestinal helminths by the species were as follows: Ascaris lumbricoides 26.3%, Echinostoma sp. 15.6%, hookworm 6.4%, Opisthorchis sp. 4.0%, Rhabditis sp. 2.4%, and Trichuris trichiura 0.4%. The infection rate of intestinal protozoa were as follows: E. coli 7.6%, G. lamblia 3.2%, I. butschlii 3.2%, and E. histolytica 0.8%. More than two different kinds of parasites were found in 16.7% of the stool samples. All the children infected were treated with albendazole, praziquantel and metronidazole according to parasite species. The results showed that intestinal parasites are highly endemic in this area.

Epidemiological study of clonorchiasis and metagonimiasis along the Geum-gang (River) in Okcheon-gun (County), Korea

The endemic status of clonorchiasis and metagonimiasis along the Geum-gang (River) in Okcheon-gun (County) in Korea was examined. From February to December 2000, stools of total 1,081 inhabitants living in 5 villages were examined. Each stool specimen was examined by both the cellophane thick smear method and the formalin-ether sedimentation technique. Egg-positive cases were further analyzed by Stoll's egg-counting technique, and praziquantel was administered to positive cases. The egg-positive rates for Clonorchis sinensis and Metagonimus species were 9.3% and 5.5%, respectively, and the double infection rate was 3.5%. The numbers of eggs per gram (EPG) of feces of C. sinensis and Metagonimus sp. were 918 +/- 1,463 and 711 +/- 947, respectively. The egg-positive rates for C. sinensis and Metagonimus sp. in the riverside area were 14.2% and 8.4%, respectively, which were significantly higher than those of the inland area (3.2% and 1.7%, respectively). The egg-positive rates of C. sinensis and Metagonimus sp. in males (16.7% and 10.0%) were significantly higher than those of females (3.5% and 1.8%). However, there were no significant differences of EPG values between localities and sexes. The prevalence of clonorchiasis and metagonimiasis in this survey was significantly lower than that in the previous reports. However, there is still a high prevalence of infection with C. sinensis and Metagonimus sp. in this region, especially in the riverside area.

A nationwide survey of the prevalence of human Gymnophalloides seoi infection on western and southern coastal islands in the Republic of Korea.

A nationwide survey was performed to know the distribution and prevalence of human Gymnophalloides seoi infection on western and southern coastal islands in the Republic of Korea. A total of 4,178 fecal specimens were collected from residents on 45 (24 western and 21 southern) islands, and examined by Kato-Katz and formalin-ether sedimentation techniques. Eggs of G. seoi were detected from 160 (3.8%) people living on 22 (13 western and 9 southern) islands. The prevalence varied by the location of islands; higher on western islands than on southern islands. The highest prevalence was found on Amtaedo (25.3%), followed by Cheungdo (25.0%), and Anchwado (20.9%) (Shinan-gun). A little lower prevalence was observed on Munyodo (13.3%), Shinshido (12.9%), and Sonyudo (10.3%) (Kunsan-shi). Of the remaining islands, the regions showing the prevalence greater than 5% included Kohado, Dallido (Mokpo-shi), Pyeongildo, Kogumdo (Wando-gun), and Keogumdo (Kohung-gun). A strong age predilection was noted (p < 0.05); 95% of the infected people were over 40 years old. Females showed a little higher prevalence than males. The results indicate that human G. seoi infection is more widely distributed than previously considered. Nine of 11 islands (excluding the 2 known areas Munyodo and Sunyudo) that showed greater prevalence than 5% are regarded as new endemic foci of G. seoi.

A small-scale survey on the status of intestinal parasite infections in rural villages in Nepal.

The status of intestinal parasite infections was investigated in two rural villages (Chitrasar, Jerona) in Chitwan District, Nepal in 1999. Stool examination was performed with a total of 300 specimens from schoolchildren by formalin-ether sedimentation technique. The prevalence rate of intestinal parasite infections in the surveyed areas was 44.0%. The prevalence rate in Jerona was slightly higher than that in Chitrasar. The prevalence rate of intestinal parasite infections in female was slightly higher than that in male without statistically significant difference. Entamoeba coli was the most commonly found protozoan parasite (21.0%) followed by Giardia lamblia (13.7%) and others (5.3%). Hookworm was the most prevalent intestinal helminth (13.0%) followed by Trichuris trichiura (3.0%) and others (5.0%). Forty-three specimens (14.3%) showed mixed infections. It is necessary to implement large-scale treatment with anthelminthics, health education and sanitary improvement for intestinal parasite control in the surveyed areas.

High endemicity of Metagonimus yokogawai infection among residents of Samchok-shi, Kangwon-do.

A small-scale epidemiological survey was undertaken during 1997-1998 on the residents along the Osib-chon (Stream), Samchok-shi (City), Kangwon-do (Province), to evaluate the status of Metagonimus yokogawai infection. A total of 165 fecal samples was collected and examined by cellophane thick smear and formalin-ether sedimentation techniques. The egg positive rate of M. yokogawai was 29.7%, showing a remarkable difference between males (46.6%) and females (16.3%). To obtain the adult flukes of M. yokogawai, 11 egg positive persons were treated with praziquantel and purged with magnesium sulfate. A total of 242,119 adult flukes (average 22,010 per person, 367-119,650 in range) was collected from diarrheic stools, all of which were identified as M. yokogawai. The results show that M. yokogawai is still highly endemic in this area.

Increased sensitivity of routine laboratory detection of Strongyloides stercoralis and hookworm by agar-plate culture

The efficacy of agar-plate culture has been evaluated for the detection of Strongyloides stercoralis and hookworm, compared with direct smear, the formalin-ether sedimentation technique and the filter-paper method. Of 1085 stool samples from the routine laboratory service at King Chulalongkom Memorial Hospital in Bangkok, 241 samples harboured S. stercoralis, 153 hookworm and 2 Rhabditis hominis. The recovery rate of S. stercoralis by agar-plate culture is significantly superior to the other methods ( P < 0·005). The ratios of positive results from the methods used to the total number of S. stercoralis-positive cases were as follows: 1:1·03 by agar-plate culture, 1:1·85 by the filter-paper method, 1:1·98 by the sedimentation technique and 1:10·48 by direct stool smear. A similar trend of the efficacy ratio of each method was obtained for hookworm detection. The characteristic furrows left by hookworm larvae, and larvae and adults of S. stercoralis could be used for preliminary species identification. Daily search for furrows on agar plates for up to 6 consecutive days resulted in an increased sensitivity for diagnosis of both S. stercoralis and hookworm infections.