Epidemiological features of visceral leishmaniasis in Fars province, southern Iran

To describe the epidemiological features of pediatric visceral leishmaniasis in southern Iran. This retrospective study was carried out using local hospital records of VL patients from 2001 through 2009. Data such as age, gender, place of residence, clinical signs and symptoms, treatment, history of recurrence were recorded. The collected data were statistically analyzed using SPSS software. A total of 260 cases of VL have been recorded during 2001 to 2009 in south of Iran, based on hospital records. Mean age of patients was 3.5 years with the highest prevalence in 2 years old patients. The diseases have been more common in males [60%]. The main clinical signs and symptoms of the patients were fever [96.2%], hepatosplenomegaly [68.8%] and abdominal protrusion [71.9%]. Most of cases were from Kazeroun County [17.5%] in Fars Province followed by Borazjan in Boushehr Province. Bone marrow aspirations have been performed in 178 of cases and Leishmania amastigotes were detected in only 50 [28.1%] cases. Glucantime has been the first drug treatment while 19.3% of cases have been treated with amphotericin B. Mortality rate was 6.2% and children under 1 year old have been the main victim of the disease. Relapse has been noted in 7.3% of patients. VL is still endemic in South of Iran, especially in Fars Province with a noticeable mortality rate. Moreover, cases of the diseases are reporting from neighboring provinces and this might be due to the spreading of the diseases to the adjacent provinces

Seroprevalence of human fasciolosis in a new-emerging focus of fasciolosis in Yasuj district, Southwest of Iran

Fasciolosis is an important health and veterinary problem in Iran. The epidemiological pattern of disease has been changed markedly in recent years and there are regions that have potent capacity to be new focus of the disease. One of these areas is Yasuj district in Southwest of Iran where animal fasciolosis has been quite common. The current study was conducted to determine the seroprevalence of human fasciolosis in this area and to reveal the epidemiological factors associated with the spreading of the disease in this region. One thousand blood samples were randomly collected from five villages in Yasuj district. ELISA, using Fasciola somatic antigen [SA], was carried out to detect anti Fasciola antibodies in the collected sera. Anti-Fasciola antibodies were detected in serum of 18[1.86%] individuals by ELISA. Out of 18 seropositive people, 9 [0.9] were female and 9 [0.9%] were male. Most of people [99.8%] had a history of consuming wild freshwater plants mainly Nasturtium microphyllum [local name Bakaloo] and/or Mentha logifolia [local name Pooneh]. No significant correlation was found between seropositivity to fasciolosis and sex, age, history of consumption of green leafy aquatic plants whereas correlation between seropositivity and abdominal pain was significant [P< 0.05]. Results of this study showed that the seroprevalence rate of human fasciolosis in Yasuj district is relatively high and this area can be considered as a new emerging focus of the disease in Iran

Comparison of agar dilution, broth dilution, cylinder plate and disk diffusion methods for evaluation of anti-leishmanial drugs on Leishmania promastigotes

Leishmaniases are a group of diseases caused by Leishmania parasites. Growing of drug unresponsiveness in leishmaniasis patients necessitates the development of new drugs and accordingly a suitable assay is needed for evaluation of any modalities. The aim of this study was to compare four drug assays methods, agar dilution, broth dilution, cylinder plate and disk diffusion, for evaluation of anti-leishmanial drugs on Leishmania promastigotes, using glucantime as a currently available drug for treatment of leishmaniasis. For broth dilution method, different concentration of glucantime was added to the parasite culture [promastigotes of Leishmania], while in cylinder plate method wells were punched in agar gel and filled with different concentration of drug and zone of inhibition was measured in each well. In disk diffusion method, the parasites were cultivated on the surface of agar; filter paper disks were enriched with various concentration of glucantime and were placed on the surface of agar. In agar dilution method, various concentrations of drug were incorporated onto blood agar and the parasites were cultivated on the surface of the agar. A direct correlation was found between the drug concentration and size of inhibitory zones in cylinder plate and disk diffusion methods. These two drug assays methods provided much better performance in comparison with broth and agar dilution methods. Cylinder plate and disk diffusion methods seem to be acceptable methods for susceptibility testing of anti-leishmanial compounds on Leishmania promastigotes

Expression and purification of P43 Toxoplasma gondii surface antigen

Toxoplasma gondii is an obligate intracellular protozoan parasite, capable of infecting all species of mammals including man. Congenital toxoplasmosis is more important during pregnancy for the first time. In this study we expressed and purified P43 Toxoplasma gondii tachyzoite and bradyzoite specific surface antigen. The recombinant pGEMEX-1 contained Toxoplasma P43 coding sequence was transformed into E. coli and mass cultured in LB medium contained 100 micro g/ml ampicillin at 37 [degree sign]C over night .The T7 promoter was induced by 1mM isopropyl-1-thio-beta-D-galactopyranoside [IPTG. Recombinant protein was purified by affinity chromatography and confirmed by gel diffusion dot blot and western blot,-using specific anti Toxoplasma antibodies. Recombinant plasmid was induced by IPTG and analyzed by SDS-PAGE. Recombinant protein was confirmed by Western-blot and dot blot using anti human Toxoplasma antibody. Recombinant Toxoplasma P43 was produced successfully

Performance of antigen B isolated from different hosts and cyst locations in diagnosis of cystic echinococcosis

The aim of this study was to assess the performance of Antigen B [AgB] isolated from different Echinococcus granulosus intermediate hosts and from different cyst locations for immunodiagnosis of human cystic echinococcosis [CE]. Hydatid cyst fluids were collected from lung and liver cysts of sheep, liver cysts of goats, lung cysts of camels and cattle, and liver cysts of human. AgB was purified from each of these hydatid cysts fluids. Serum samples obtained from 47 pathologically confirmed cases of CE along with 30 sera samples from non-CE patients and 40 sera from healthy individuals were tested by ELISA using AgB prepared from different hosts or cyst locations. The highest sensitivity [97.8%] for diagnosis of CE was seen with AgB prepared from human liver cysts. This maximal sensitivity was followed by AgB isolated from those of sheep liver and lung cysts. The least sensitivity was found with AgB prepared from bovine lung cysts. The highest specificities [97.1%] were observed with AgB isolated from human liver cysts followed by those of sheep and goat liver cysts while the lowest specificity was seen with AgB isolated from bovine lung cysts. In view of the specificities and sensitivities of the different AgB, the best validity was found for AgB prepared from human liver cysts while the least validity was found with AgB prepared from bovine lung cysts. For any AgB-based tests, obtaining of the antigen from one of these sources will significantly increase the diagnostic sensitivity and specificity of the assay

[Effect of echinacea purpurea on control of leishmania major induced cutaneous leishmaniasis in mice]

Leishmaniasis is a disease caused by intracellular protozoa parasites of the genus Leishmania and is endemic in some areas of Iran. Echinacea purpurea is a native plant from North America which is one of the most important medical herbs known with immuno-stimulant properties. This study was performed to determine the effect of alcoholic extract of Echinacea purpurea on prophylaxis and treatment of Leishmania cutaneous lesions. In this experimental study which was conducted at Shiraz University of Medical Sciences in 2009, eighteen mice were divided into 3 groups. Group one received Echinacea purpurea extract [200 mg/ml] in their water, for 2 weeks before parasite injection, while group two were first injected with parasite amastigotes, followed by administration of Echinacea purpurea extract for 2 weeks. Group three was the control group, which received parasites, but not the extract. The size of Leishmania lesions in the tail base, right and left foot were measured with vernier caliper. The lesion areas were calculated and the collected data were analyzed with SPSS software. The mean of lesion size in each group of mice were compared and analyzed. No significant differences in the lesions size were found between the three mice groups. Therefore, Echinacea purpurea extract was not effective against Leishmania major based on the findings of this study. Our findings suggest that Echinacea extract is not effective in treatment or prophylaxis of leishmaniasis in mice. Yet, further studies are needed to determine the effects of other extracts of this plant

Seroepidemiological study of visceral leishmaniasis in Booyerahmad district, south-west Islamic Republic of Iran

Visceral leishmaniasis [VL] is endemic in parts of Islamic Republic of Iran. A cross-sectional seroprevalence study was carried out in children in Booyerahmad district in the south-west of the country. Serum samples were taken from 1628 children up to 10 years old from different areas in Booyerahmad in 2005-06. Anti-Leishmania antibody was detected in 50 out of 1628 children [3.1%] by direct agglutination test [antibody titre > 1:3200]. There was no significant difference in seropositivity between the sexes [2.8% males and 3.3% females]. The highest rate of infection [5.2%] was in the age group 10 years. Further studies are needed to explore the reservoirs and vectors of the disease in this region

[Seroprevalence of HIV infection among high-risk groups in Kohgiloyeh and Boyerahmad province in 2009]

According to WHO report, there are more than 40.3 million people infected with HIV worldwide. Detection of HIV-infected people in each community helps to control and prevent the infection. This study aimed to evaluate the prevalence of HIV infection among high-risk groups in Kohgiloyeh and Boyerahmad province in Iran. This was a cross-sectional study which was conducted in 2009 in Kohgiloyeh and Boyerahmad province in Iran. High-risk groups for HIV were the subjects of this study. Blood samples were taken from 2009 from 2009 people from Boyerahmad Gachsaran, and Dehdasht and tested by ELISA for anti-HIV 1/2 antibodies. Demographic features of participants were recorded using a questionnaire during sample collecting. Collected data were analyzed by SPSS version 13 software. From 2009 subjects, HIV antibody was detected in 36 [1.8%] of cases. High prevalence of infection [3.2%] was seen in 31-40 years age group. The rate of infection was higher in males compared to females. Unemployed people were found to be the main victim of the disease. The highest rate of infection was seen in Gachsaran in comparison to other townships. Findings of the study demonstrated that HIV infection is prevalent in high-risk groups in Kohgiloyeh and Boyerahmad province and control measurements should be implemented to prevent the distribution of the infection

Comparison of three methods for diagnosis of cutaneous leishmaniasis

Leishmaniasis is one of the infectious parasitic diseases of highest incidence in the world. Cutaneous Leishmaniasis [CL] has long been reported in Shiraz, Southern Iran. There is a need to find a sensitive and specific method for treatment and control of the disease. We have compared the sensitivity of the conventional methods microscopy and cultivation of lesion scrapes against PCR amplification of parasite kinetoplast DNA from these samples. The samples [n=219] were obtained from the patients clinically suspected of CL. The smears were stained with Giemsa for microscopy and cultured in Novy-Nicolle-McNeal [NNN] blood agar for promastigote growth. For PCR, the dry smears were scraped off the slides and DNA was extracted. The positive rates from 219 specimens were 76.71%, 50.68%, and 93.61% for microscopy, cultivation, and PCR, respectively. The highest correlation was found between PCR and microscopy method [P= 0.014]. In PCR assay, 95.61%, 3.9%, and 0.49% of the samples were identified as Leishmania major, L.tropica, and dermatropic L.infantum, respectively. The PCR method appears to be the most sensitive for the diagnosis of CL and is valuable for identifying the other species of Leishmania with confusing dermatropic signs

In vitro infectivity of leishmania major isolated from patients with different clinical forms of cutaneous leishmaniasis and its association with parasite zymodems

The aim of this study was to characterize the Leishmania parasites isolated from cutaneous leishmaniasis [CL] patients in Pars Province in Iran and to compare the potential infectivity of the isolates in macrophage cell line. Moreover, attempt was made to find out the association between parasite infectivity and their zymodems. Twenty samples were taken from the skin lesion of CL patients. The samples were cultured in biphasic media followed by mass cultivation in RPMI medium. Each isolate was tested for the activity of the 5 enzymes including glucose phosphate isomerase [GPI], malate dehydrogenase [MDH], nucleoside hydrolase 1 and 2 [NH1 and NH2], and phosphoglucomutase [PGM]. The enzymatic profiles of the isolates were compared with WHO reference strains. Specific PCR [primers: LIN17 and LIN R4] and RAPD-PCR were used as complementary methods for characterization of the isolates. Isoenzyme electrophoresis showed that all of the isolates were L. major. PCR with LIN 17 and LIN R4 and RAPD-PCR with AB-07 primers further determined the isolates as L. major. Results of macrophage infectivity experiment, using J774 cell line, showed that the most virulent isolates were related to Z1 with 63% macrophage infectivity rate. A well correlation was found between the infectivity rate of the isolates and type of ulcer. Those isolates with high infectivity rate were involved in more severe, ulcerative or erythmatose lesions in CL patients. The most invasive isolates might be a good candidate for immunological studies and for vaccine development

[Prevalence of hepatitis B antibodies in health-care workers in Yasuj Hospitals]

Hepatitis B is a common infection in the world and one of the main health problems in our country. Over 350 million people are infected with Hepatitis B virus in the world and are chronic carriers of this infection. Health care workers are at risk of infection with blood born viruses including hepatitis B [HBV]. This study was conducted to find out the rate of anti-HBs antibodies among the health-care workers [HCW] in Yasuj hospitals, Southwest of Iran. This is a cross sectional descriptive study in which 212 staff was randomly selected from different wards of the hospitals in Yasuj. Blood samples were taken from each individual and tested for hepatitis B surface antibody [anti-HBs] by ELISA. Those who had anti-HBs titer > 10 IU/ml were considered as positive. Collected data were analyzed by SPSS software using descriptive data analysis and chi-square test. 61.3% of the subjects were female and 38.7% were male. 93.9% of the subjects had a history of one to three doses of hepatitis B vaccination. Results of this study showed that 185 [87.3%] of the staff have anti hepatitis B antibodies [Anti-HBs]. Among the staff that was negative for anti-HBs antibody, 12 had a history of hepatitis B vaccination [at least one dose]. Female employees were more positive than males [93% vs. 78%] and this difference was statistically significant [p<0.05]. Moreover, a positive correlation was found between the titer of antibody and sex where females had a higher titer of antibody in comparison with males [p<0.05]. No correlation was found between the workplace of HCW and positive anti-HBS. Result of this study indicates that more than 85 percent of the health-care workers in Yasuj have reasonable immunity against hepatitis B infection. A small proportion of HCWs had no immunity against HBV. The second course of hepatitis B vaccine should be delivered to those who had no immunity against hepatitis B despite their receiving the first course of vaccination

[Serological study of visceral leishmaniasis in Boyer Ahmad Township in 2005]

Visceral leishmaniasis [Kala-azar] is a disease caused by intracellular protozoa parasites of the genus Leishmania. Kala-azar in Iran is usually seen in children below 10 years old. Kala-azar is endemic in some areas of Iran including Ardebil, Fars, and East Azerbaijan. The present study was designed to assess the situation of Kala-azar in children of ten and below ten years in Boyer Ahmad township in Kohgiloyeh and Boyer Ahmad province in 1384. Blood samples were collected from all the children of 10 and less than 10 years old. In this survey 1628 blood samples were collected from finger tip of children on filter paper. Collected samples were evaluated by direct agglutination test [DAT]. Collected data were analyzed by SPSS software. Result of this study showed that from 1628 samples collected, 50 cases [3.07 percent] were become positive with DAT test. Infection in females was less than male. No significant correlation was found between antibody titer and sex. The highest prevalence rate of the infection was seen in age 10 [14.82%]. Leishmania Infection was not found in any of the studied reservoirs [dogs]. Results of this study indicate that Kala-azar is an endemic disease in the studied region. Considering the obtained data, more study is needed to find out the prevalence of the disease in other regions of the province and also to find ou more about the possible animal reservoir of the diseases in the region

Application of western blotting using native antigen B for serodiagnosis of human cystic echinococcosis

Hydatidosis is one of the most important helminthic diseases causing serious health and economic problems in many countries including Iran. Currently available diagnostic approaches for diagnosis of cystic echinococcosis [CE] are not satisfactory since they have not achieved a reasonable validity in diagnosis of CE. This study aimed to assess the performances of Western blotting [WB], using native antigen B, for serological diagnosis of human hydatidosis. Antigen B was prepared from sheep hydatid cyst fluid. Serum samples obtained from 40 surgically proven cases of hydatidosis along with serum samples from non-hydatidosis patients and samples from healthy persons were tested by immunoblotting, using native antigen B. From 40 sera of hydatidosis patients, 32 cases [80%] detected 8 kDa subunit, 29 cases [72.5%] recognized the 16 kDa component and 29 cases [72.5%] detected 24 kDa subunit of antigen B. The highest sensitivity [80%] was achieved with 8 kDa subunit of antigen B while the other components [16 and 24 kDa] showed a lower sensitivity [72.5%]. Samples from healthy controls and non-hydatidosis patients did not yield any detectable band in the Western blotting assay. Accordingly, specificity of the system was found to be 100%. Considering the use of native antigen B, application of western blotting, based on 8 kDa subunit of antigen B, is highly recommended for the confirmatory diagnosis of hydatidosis

Direct agglutination test and enzyme linked immunosorbent assay with urine samples for the diagnosis of visceral leishmaniasis

Visceral leishmaniasis [VL] or Kala azar is an infectious disease caused by various species of Leishmania parasites. The aim of this study was to detect and compare the presence of anti-Leishmania antibodies in the urine of visceral leishmaniasis patients using ELISA and DAT methods. A total of 30 urine samples were collected from VL patients referred to Shiraz [southeast of Iran] hospitals. Moreover 31 urine samples were collected from healthy individuals and patients with other diseases such as malaria, brucellosis, hydatidosis and cutaneous leishmaniasis. Collected samples were examined to detect anti-Leishmania antibodies in urine, using ELISA and DAT. Anti-Leishmania antibody was detected in urine of 18 out of 30 [60%] VL patients by DAT while ELISA detected anti-Leishmania antibodies in urine of 28 out of 30 [93.3%] of VL cases. Sensitivity and specificity of urine-based DAT was 60% and 83.9%, respectively while sensitivity and specificity of urine-based ELISA were 93.3% and 93.5%, correspondingly. Urine-based DAT and ELISA have a reasonable specificity and sensitivity in diagnosis of VL. Accordingly, urine-based ELISA might be a suitable alternative for serum based assays for diagnosis of VL

[Human cystic echinococcosis in Yasuj: a survey of ten year hospital records]

Cystic echinococcosis [CE] is a zoonotic infection of humans and domestic animals caused by larvae of the cestode Echinococcus granulosus. Echinococcosis has a world-wide geographic distribution and occurs in all continents. CE is one of the most important zoonotic diseases prevalent in different parts of Iran and human cases are repeatedly reported from medical centers in different regions of Iran. This study was conducted to survey the hospital records of hydatidosis between 1996 and 2006 in Yasuj, southwest of Iran. Hospital records of CE patients from 1996-2006 in three hospitals in Yasuj were carefully studied. Data such as age, sex, occupation, place of residence, number of cyst, cyst location, clinical signs and other related features were recorded in a predestined information sheet. Collected data were statistically analyzed. Results of this study showed that during 1996 to 2006 [10 years] 105 cases of hydatidosis have been admitted in Yasuj hospitals, out of which 70 cases [66.7%] were women and 35 cases [33.3%] were men. Considering the age of patients, the highest rate of infection was recorded in 31-40 year old patients. Female housekeepers with 66.7%of cases were found to be the main victims of the disease. Hepatic cyst was recorded in 81% of cases where nephritic cyst was the second most prevalent one. Most of the patients [87.8%] has only one cyst, though 11.4% of the patients had 2 cysts and in one case 18 cysts have been isolated from the patient. Recurrence of the disease, based of number of patient's operation, was noted in 14.3% of cases and in one case five times of surgery for removing of hydatid cyst were noted. Results of this study indicate that CE is an endemic disease in the studied area. Data also suggest tat the trend and number of CE cases remained unchanged during the 10 years in this area. Therefore, effective steps should be taken to control the disease in this hydatid cyst endemic district

[Characterization of leishmania parasites isolated from kala- azar patients in kohgiloyeh and boyerahmad, using semi-nested PCR]

Visceral leishmaniasis [VL] is a disease commonly known as Kala-azar caused by protozoan parasites of the genus Leishmania including L. donovani, L. infantum and L. chagasi. VL is sporadic in manyareas of Iran and is endemic in a few provinces such as Fars, Azarbayjan, Bushehr, Ardabil and Qom. VL has been reported from some areas of Kohgiloyeh and Boyerahmad and this study aimed to characterize the causative agent of VL in this region. Bone marrow sample was obtained from 6 VL patients from children department in Imam Sajad hospital in Yasuj. DNA was extracted from the obtained samples and was checked by semi-nested PCR to determine the species of the parasite. To do that, a segment of minicircle kinetoplast DNA was amplified, using LINR4 and LIN17 primers. Products of PCR were evaluated by electrophoresis, using 1.5% agarose and stained with ethidium bromide. Parasitologically examination of bone marrow smears demonstrated amastigotes form of the parasite in the samples. For mass cultivation, isolated parasites were cultured in diphasic NNN followed by RPMI 1640 media. All the samples produced a 720 bp band in PCR assay. The isolates were compared with referent strains and it was revealed that all the isolates were L. infantum. Findings of this study demonstrated that the causative agent of VL in Kohgiloyeh and Boyerahmad was L. infantum. Further study is needed to explore other aspects of VL in this region