Toxocara Canis Increases the Potential of Breast Cancer by Reducing the Expression of the P53 Protein.

INTRODUCTION: Breast cancer is considered the most frequent type of cancer in women with high mortality worldwide, and most importantly, it is the second most common cancer. However, some breast cancer-related risk factors remain unknown. So, the current study was designed to evaluate the effect of Toxocara canis on the biomarkers correlated with proliferation, apoptosis, inflammation, and angiogenesis in 4T1 tumor-bearing mice infected with Toxocara canis for the first time. METHODS: Mice were categorized into four groups: A) control, B) treated with 4T1+ Toxocara canis, C) treated with Toxocara canis, and D) treated with 4T1. The expression of Ki-67 and P53 was then evaluated by using the immunohistochemical technique. In addition, the levels of transforming growth factor-ß, Interferon gamma-γ, Interleukin 10, tumor necrosis factor-α and vascular endothelial growth factor as well as anti- Toxocara canis IgG were determined using the enzyme-linked immunosorbent assay method. RESULTS: The expression of Ki-67 was significantly increased in the 4T1+ Toxocara canis group than control and Toxocara canis groups (P < 0.001 and P < 0.001, respectively). Moreover, a significant decrease in P53 was found in the 4T1+ Toxocara canis group than in the control and Toxocara canis groups (P < 0.001 and P < 0.001, respectively). Also, the 4T1+ Toxocara canis group significantly reduced the expression of P53 more than 4T1 tumor-bearing mice (P = 0.005). In addition, the 4T1+ Toxocara canis group had an increasing tumor necrosis factor-α and vascular endothelial growth factor than controls (P = 0.004 and P = 0.002, respectively). Furthermore, a significant reduction in Interleukin 10 was found in the 4T1+ Toxocara canis group than in the control group (P = 0.004). CONCLUSION: Our findings showed that Toxocara canis could probably increase the potential of breast cancer by reducing P53 in 4T1 tumor-bearing mice infected with Toxocara canis more than other groups.

Visceral leishmaniasis-human immunodeficiency virus coinfection in a 52-year-old male in southwest Iran: a case report.

BACKGROUND: Leishmaniasis is a rare infectious disease observed in subtropical and tropical areas. This disease that demonstrates different clinical characteristics is caused by intracellular Leishmania protozoan. One of the important countries facing the incidence of this infectious disease is Iran. Recently, human immunodeficiency virus-Leishmania coinfection has been indicated in Iran. CASE PRESENTATION: In the present case report, we show an atypical case of severe visceral leishmaniasis in a 52-year-old Iranian-Arab male with positive human immunodeficiency virus status. Leishmaniasis was detected by node biopsy and subsequently histopathology evaluations and confirmed by molecular methods. CONCLUSIONS: The current study was the first report of an atypical case of a patient with Leishmania-human immunodeficiency virus coinfection in southwestern Iran, which was not responsive to the treatment. Therefore, the health authorities should be aware of these reports, which require permanent clinical follow-up of the patients as well as effective treatments.

Molecular Identification of Free-living Amoebae Isolated from Hospital Water Systems in Southwest Iran.

INTRODUCTION: Free-living amoebae are opportunistic amoebae that usually live freely in various environmental conditions, including warm water and even in water supply network pipes and reservoirs connected to water. In addition to living freely, these protozoa are able to attack the host's body when they enter a human or animal body. Therefore, it is necessary to identify their presence in water resources. So, this study aimed to identify free-living amoebae isolated from water reservoirs of hospitals in southwest Iran. METHODS: A total of 80 water samples were isolated from the hospitals of Ahvaz city, southwest Iran, and their physical and chemical parameters were measured. The samples were then put into non-food agar culture medium and stained using the Wright-Giemsa staining. Finally, the samples were identified by the PCR molecular method. RESULTS: The mean pH and turbidity values were 7.57 ± 0.03 and 3.31 ± 0.26 nephelometric turbidity unit (NTU), respectively. The mean residual chlorine and electrical conductivity were 0.91 ± 0.02 and 1122.39 ± 24.31, respectively. In addition, 9 (11.25%) and 3 (3.75%) samples were contaminated with Acanthamoeba spp. and Naegleria spp., respectively. However, no positive cases of Balamuthia spp. infection were observed. Moreover, two samples were co-infected with Acanthamoeba spp. and Naegleria spp. CONCLUSION: Due to the existence of free-living amoebae in water storage tanks at hospitals, it is necessary to prevent possible contamination with these amoebae and infectious agents by using new methods of disinfection and purification of water resources.

Is Immuno-PCR Better than ELISA Test for Detection of Toxoplasma gondii IgG Antibody?

INTRODUCTION: IgG antibodies against T. gondii persist for years, and can act as a reliable serological biomarker for the diagnosis of previous exposure to this parasite. Hence, the current investigation was designed to compare diagnostic power of immuno-polymerase chain reaction (iPCR) and enzyme-linked immunosorbent assay (ELISA) methods for detection of T. gondii IgG antibody. METHODS: Immunoglobulin M (IgM) and immunoglobulin G (IgG) antibodies against T. gondii were measured by the ELISA method in 81 participants. In addition, detection of acute and chronic toxoplasmosis was performed via the ELISA IgG avidity. The set-up of iPCR was carried out and then, serum IgG of subjects were detected using the iPCR method. RESULTS: Of 81 samples, 4 (4.9%) and 30 (37%) cases were be found positive for IgM and IgG against T. gondii in the ELISA method, respectively. Moreover, of 81 specimens, 42 (51.9%) and 39 (48.1%) samples had low-avidity IgG and high-avidity IgG by the IgG avidity kit, respectively. While, 59 (72.8%) of 81 samples were detected positive using the iPCR technique. Kappa (κ) value coefficient, between the iPCR and ELISA (for IgG) showed a strong agreement (0.360, p value < 0.001). A value of 0.25 I.U./ml for serum IgG [area under curve (AUC) = 0.720 (CI = 0.613-0.827); p = 0.002] was the cut-off value to differentiating between positive and negative toxoplasmosis (with sensitivity 66.0% and specificity 60.0%). CONCLUSION: Our findings indicated despite a strong agreement shown between iPCR and ELISA methods, the diagnostic power of iPCR technique was more sensitive than ELISA test for detection of T. gondii IgG antibody. However, more complementary investigations are widely needed in this regard.

Toxoplasmosis in Treatment-Resistant Obsessive-Compulsive Disorder Patients.

BACKGROUND AND OBJECTIVES: The current study was done to evaluate the relationship between T. gondii and obsessive-compulsive disorder (OCD) as well as prevalence rate of toxoplasmosis in treatment-resistant patients with OCD in comparison with treatment-sensitive patients with OCD. METHODS: A total of 180 subjects were selected, including 90 patients with OCD and 90 control participants. Immunoglobulin G (IgG) and immunoglobulin M (IgM) antibodies against T. gondii were measured by the enzyme-linked immunosorbent assay (ELISA) technique. Detection of acute and chronic toxoplasmosis was carried out using the ELISA IgG avidity. In addition, the presence of this parasite in blood was detected using the nested-polymerase chain reaction (PCR) method. RESULTS: Regarding T. gondii IgG antibodies 42 (46.7%) and 17 (18.9%) cases were detected in the patients and controls, respectively (P = 0.000). Also, 16 (17.8%) and 5 (5.6%) cases were positive for B1 gene in patients and controls, respectively (P = 0.018). The antibodies were found to be related to risk of OCD [OR (95% CI) = 3.71 (1.88-7.30); P < 0.001]. Moreover, out of 90 patients, 35 and 55 cases were resistant and sensitive to treatment, respectively, so that 24 (68.6%) out of 35 and 18 (32.7%) out of 55 were positive for the antibodies (P = 0.01) as well as 11 (31.4%) out of 35 and 5 (9.1%) out of 55 were positive for B1 gene (P = 0.010). The antibodies were also associated with risk of resistance to treatment in patients with OCD [OR (95% CI) = 3.81 (1.42-10.17); P = 0.008]. CONCLUSION: Our findings showed that toxoplasmosis was more frequent in patients with OCD than the control group. In addition, prevalence rate of toxoplasmosis in treatment-resistant patients with OCD was significantly more than that in treatment-sensitive patients with OCD.

Genetic diversity of Leishmania major strains isolated from Phlebotomus papatasi and Phlebotomus alexandri in western Iran based on minicircle kDNA.

This study aimed to screen the natural infection rate of Leishmania major in Phlebotomus papatasi and Phlebotomus alexandri in two counties (Mehran and Dehloran) of Ilam province as cutaneous leishmaniasis endemic areas in the west of Iran. Furthermore, the genetic diversity of parasite species that are isolated from vectors, was investigated. Sandflies were collected by sticky traps from May 2018 to October 2018. Afterward, specimens were prepared for species identification by morphological features. DNA was extracted from female sandflies, and minicircle kDNA was used to identify Leishmania isolates through nested-PCR, followed by genetic diversity between Leishmania isolates was investigated by sequence analysis of the amplified minicircle kDNA. Natural infection of the L. major was shown in all positive specimens using nested-PCR. Analysis of data from 14 isolates displayed a high level of genetic diversity in L. major. In the phylogenetic trees, all of the L. major isolates occurred in six clusters. Clusters I, II, III, and VI contained isolated strains from P. papatasi. While clusters IV and V contained isolated strains from P. alexandri. Genetic diversity of L. major isolated from vectors was investigated in western Iran for the first time. According to the results of this study, probably "various clones of L. major populations are distributed in the study area.

A comparison of the efficiency of molecular and conventional methods in the diagnosis of cutaneous leishmaniosis.

Leishmaniosis is one of the most important vector borne diseases. Among different forms of the disease, cutaneous leishmaniosis (CL) is the most common. Determining the method of definitive diagnosis for the disease has been the aim of various studies. Therefore this study afforded an opportunity to investigate this subject. To diagnose CL in 150 suspected patients referred to Mehran and Dehloran health centers during June 2018 to November 2019, two polymerase chain reaction (PCR) methods were performed and compared with the in vitro culture and microscopic evaluation of stained slides. The smears were stained with Giemsa for microscopy and cultured in Novy-Nicolle-McNeal (NNN) blood agar for promastigote growth. For semi-nested PCR and PCR-RFLP, the tissue and serosity from the lesions were used for DNA extraction. The semi-nested PCR technique using minicircle kDNA gene showed the highest positivity rates among all diagnostic assays with 114/150 (76%) of the samples and was used as reference standard, followed by the PCR-RFLP test using ITS1 gene with 112/150, (74.7%) positivity rates, microscopy with 101/150 (67.3%) and then culture 72/150 (48%). microscopy and culture methods together improved overall positivity rates to 68.7% (103/150). The all positive samples using molecular technique were identified as Leishmania major. The highest sensitivity (98.3%), specificity (100%), accuracy (98.8%), negative predictive value (94.7%) and κ coefficient (0.96=almost perfect) was observed by comparing PCR-RFLP and semi-nested PCR. kDNA-semi-nested PCR and ITS1-PCR-RFLP presented an interesting alternative to conventional methods for the identification of CL and improved its diagnostic value significantly in suspected patients with negative direct smears.

The Immunization of Protoscolices P29 DNA Vaccine on Experimental Cystic Echinococosis in Balb/c Mice.

PURPOSE: Cystic Echinococosis is one of the important parasitic diseases that is considered as a problem economics and health in many parts of the world. Many efforts have been performed for controlling the disease in the world. To reach a reliable vaccine against Cystic Echinococosis is one of the important duty of governments. Several antigen of hydatid cyst for vaccine candidate have been evaluated. In this study, P-29 antigen has been used for this purpose. METHODS: E.g P29 antigen was cloned in Escherichia coli and transfected into the Chinese hamster ovary cell for antigen proliferation and used for vaccination in Balb/c mice. The recombinant antigen E.g-29 was shown using Western blot test. Two dilution of DNA vaccine (pCEgP-29) including 50 µg/100 µl and 100 µg/100 µl were prepared. Twenty four Balb/C male 6-8 week mouse were divided in 4 groups. The groups were included in 2 vaccination groups (pcEg.P29 50 µg/100 µl and 100 µg/100 µl dilution) as immunized groups and 2 groups of plasmid and PBS as control. The mice were injected intramuscularly 3 times with 2 weeks interval. After 3 weeks from last injection, all groups were challenged intraperitonealy with 2000 protoscolices. After 5 months, the mice were euthanized by ketamine/xylasine injection and number, size, and weight of cysts were recorded. RESULTS: Immunization rate was up to 93% in vaccinated group when compared with the control group. CONCLUSION: The results of this study showed that rEg.P29 could be considered as an effective vaccine for controlling of E. granulosus prevalence in intermediated host.

Study of fauna, activity patterns and Leishmania infection rate of phlebotomine sand flies in Western Iran.

Cutaneous leishmaniasis is a crucial vector-borne disease caused by various species of Leishmania and is transmitted by several species of sandflies. The present study was conducted to describe sand fly fauna on vectors of leishmaniasis and performing molecular identification of Leishmania isolates from them on the Iran-Iraq border. Entomological surveys were done from May to October 2016-2018 in 2 counties (Mehran and Dehloran) of Ilam province, west of Iran. Sandflies were collected by 40 Sticky Traps at each station. Samples were mounted for species identification using morphological characters of the head and abdominal terminalia. DNA was extracted from Phlebotomus papatasi females, and Leishmania isolates were identified through PCR on minicircle kDNA, followed by sequencing. A total of 5592 sandflies including 2 genera of Phlebotomus and Sergentomyia comprising 8 species of sand flies were detected. Leishmania major infection was detected in 3.33% of 300 tested female sandflies. Phlebotomus papatasi was predominant in outdoor and indoor resting places. Phlebotomus papatasi was determined as dominant vector of Leishmania major infection in Mehran and Dehloran counties, West of Iran. It seems the composition of sandfly species in the study area is almost similar to the other parts of Iran. A detailed description of the epidemiology and ecology of Phlebotomine sand flies needs to be established to accomplish effective vector control programs.

PCR Detection and Sequencing of Trichomonas vaginalis in Women with Suspected Vaginitis in Southwestern Iran.

INTRODUCTION: The human being is considered a natural host for Trichomonas vaginalis (T. vaginalis), which causes trichomoniasis, the most frequent non-viral sexually transmitted infectious disease in the world. The current study aimed to evaluate the prevalence and sequencing of T. vaginalis in women with vaginitis. ; Methods: In the current research, 514 vaginal discharge samples were obtained from women with vaginitis. The specimens were evaluated by the direct wet mount examination, Dorset culture medium, and PCR technique. Primers were designed for the detection of TVK3/TVK7, TVA5/TVA6 genes specific for the identification of T. vaginalis. The PCR-positive samples were sequenced and compared with the sequences registered in the GenBank database. ; Results: Among the collected samples, 30 (5.83%), 45 (8.75%), 90 (17.50%), and 62 (12.06%) cases were positive for T. vaginalis when assayed by the direct wet mount examination, Dorset culture medium, and PCR technique (TVK3/TVK7, TVA5/TVA6 genes), respectively. There was no significant relationship between trichomoniasis and demographic characteristics of women, such as age, occupational status, mode of delivery, number of deliveries, educational level, and contraceptive methods (p˃0.05). The range of vaginal pH was between 5-7 in women with vaginitis, and there was a significant statistical correlation between the pH values and the infection rate (p<0.05). The PCR-positive samples had 100% sequence homology with the reference sequence in the GenBank database (accession number L23861.1). ; Conclusion: This study confirmed a relatively high prevalence of T. vaginalis in the southwestern region of Iran. According to our results, the PCR method, especially detecting TVK3/TVK7 genes, was more sensitive than the direct wet mount examination and Dorset culture medium methods.

Comparison of the Detection of Toxocara Spp. in the Soils of Public Parks of Ahvaz (Southwest of Iran) by PCR and Loop-Mediated Isothermal Amplification (LAMP).

INTRODUCTION: The infections caused by Toxocara spp. are considered as one of the most important zoonotic diseases in the world, especially in developing countries. Human toxocariasis, particularly in children, is acquired by playing in public parks. Hence, the aim of the current study was to detect Toxocara spp. in the soils of public parks of the city of Ahvaz, southwest of Iran, using the PCR and loop-mediated isothermal amplification (LAMP) methods. METHODS: In this descriptive cross-sectional study, 260 soil samples were randomly collected from the different public parks of the city of Ahvaz. After performing zinc sulfate (ZnSO4) flotation technique, the DNA samples were extracted from the isolated Toxocara spp. eggs. Lastly, the extracted DNA was used for PCR and LAMP-based molecular detection. RESULTS: Out of 260 specimens, 57 (21.9%) samples were found positive for Toxocara spp., using the PCR method, out of that 38 (14.6%) samples were positive for T. canis and 19 (7.3%) samples were positive for T. cati. Also, out of 260 specimens, 81 (31.1%) cases were positive for Toxocara species, using the LAMP method, among them 51 (19.6%) samples were found positive for T. canis and 30 (11.5%) samples were positive for T. cati. Kappa (κ) coefficient between PCR and LAMP showed a strong agreement (0.766, P-value=0.002). CONCLUSION: The obtained data showed a relatively high outbreak of Toxocara spp. in the public parks' soils of the city, using the PCR and LAMP methods. Since the parasite can cause human toxocariasis, particularly in children; thus, the health authorities of the city of Ahvaz and other similar cities, especially in developing countries, must pay more attention to the hygiene of the public parks' soils.

Evaluation of immunogenicity and protective effect of DNA vaccine encoding surface antigen1 (SAG1) of Toxoplasma gondii and TLR-5 ligand as a genetic adjuvant against acute toxoplasmosis in BALB/c mice.

AIMS: Toxoplasma gondii is an obligate intracellular, protozoan that causes a high incidence of serious zoonotic parasitic disease in humans. In the present study the immune-protective efficacy of a DNA vaccine encoding SAG1 in combination with a gene sequence encoding FliC of Salmonella typhimurium (Toll-like receptor 5 agonist) was evaluated against acute T. gondii infection in mice. METHODS AND RESULTS: Ninety-nine female inbred BALB/c mice were divided into nine groups of 11 mice and were immunized intramuscularly three times at three-week intervals (days 0, 21 and 42) and challenged with virulent T. gondii RH strain 4 weeks later. The immunization of pVAX1-SAG1 administered with pVAX1-fliC in mice indicated specific humoral responses, with higher IgG antibody titers and a mixed IgG1/IgG2a response than in other groups (with a predominance of IgG2a over IgG2b and IgG1). Also, the cellular immune response elicited high levels of IFN-γ and IL-12 cytokines and low levels of IL-4 production compared to traditional adjuvants. Furthermore, the mice vaccinated with pVAX1-SAG1+pVAX1-fliC survived for slightly longer after the last immunization and challenge with the T. gondii. CONCLUSION: This investigation indicated that cocktail DNA vaccine encoded SAG1 gene of T. gondii and FliC can protect against acute toxoplasmosis.

Higher prevalence of Blastocystis hominis in healthy individuals than patients with gastrointestinal symptoms from Ahvaz, southwestern Iran.

BACKGROUND: Blastocystis, a common intestinal protozoan of humans and animals, infected more than 1 billion people around the world. This enteric protozoan is frequently reported in both healthy individuals and patients with gastrointestinal (GI) symptoms. METHODS: Three hundred and forty-five fecal samples including 151 GI patients and 194 healthy individuals were examined by microscopy, culture and PCR-sequencing techniques to determine Blastocystis frequency and subtype (ST) variation. RESULTS: The occurrence of Blastocystis was detected 56 (16.2%) and 85 (24.6%) by microscopy, culture and PCR methods, respectively. Out of the 85 positive patients, 60 (70.6%) were asymptomatic and 25 (29.4%) were symptomatic. The results of 41 successfully sequenced isolates identified 8 (19.5%), 8 (19.5%), and 25 (61.0%) ST1, ST2, and ST3, respectively. CONCLUSION: This study has found that Blastocystis was more common in healthy individuals than GI patients. Another finding was that no correlation was found between clinical symptoms and Blastocystis STs.

ERRATUM - Paleoparasitology in Iran: A Review.

Erratum Following publication of the Original article "Paleoparasitology in Iran: a review" (Infez. Med., volume 26, issue 4, pages 364-402, year 2018). We became aware that the correct affiliation 3) is: Health Research Institute, Infectious and Tropical Diseases Research Center, Ahvaz Jundishapur University of Medical Sciences, Ahvaz, Iran and that the acknowledgements were included by mistake in the original paper and have to be considered as deleted.

Molecular Genotyping of Giardia duodenalis in Humans in the Andimeshk County, Southwestern Iran.

INTRODUCTION: Giardia duodenalis, one of the most common intestinal protozoa, infects a wide range of vertebrates, including humans and animals. MATERIALS AND METHODS: In this study, 84 Giardia duodenalis positive stool samples were collected from 3580 patients attending the Imam Ali Hospital and two public health centers in Andimeshk County, southwestern Iran. Stool samples were examined initially by microscopy, and then G. duodenalis was confirmed by SSU rRNA gene and genotypes were determined by amplification of the gdh and ß-giardin genes. RESULTS: The SSU rRNA, gdh, and ß-giardin genes were successfully amplified in 89.3%, 58.3%, and 51.2% samples, respectively. Of the positive samples for gdh and ß-giardin, 40 isolates were successfully sequenced. Twenty-three isolates belonged to assemblage A, sub-assemblage AII, and 17 belonged to assemblage B. Of the 24 successfully amplified asymptomatic cases, 12 belonged to assemblage A and 12 belonged to assemblage B. CONCLUSION: The current study found that 64.3% of the patients were asymptomatic. From an epidemiological point of view, the high percentage of asymptomatic patients is important because of their role in the transmission of Giardia. The predominant assemblage was assemblage A, sub-assemblage AII. In general, therefore, it seems that most infections are probably transmitted by anthroponotic pathways in the region.

Screening municipal waste collectors for cystic echinococcosis and toxocariasis in southwestern Iran.

INTRODUCTION: Cystic Echinococcosis (CE) and toxocariasis caused by the larval stages of intestinal dog worms including Echinococcus granulosus and Toxocara spp. are among the most widespread zoonotic diseases. METHODOLOGY: Four hundred municipal waste collectors were serologically evaluated for CE and toxocariasis. To identify the seropositive cases of CE, an ELISA test was performed using native AgB. Toxocara IgG detection was carried out using ELISA DRG kit (USA), and the seropositive cases were then examined by a Western blot kit (LDBIO, France) to confirm the positive ELISA results. RESULTS: 15 (3.7%) workers were seropositive for CE according to the ELISA. A significant relationship was observed between being seropositive and having contact with soil and dogs. No significant correlations were observed between education and the prevalence of these diseases. Of the 15 seropositive workers for CE, ten worked in district 5 of Ahvaz. Toxocara IgG was identified in 11 (2.7%) cases using the ELISA; however, none of them were confirmed by Western blotting. CONCLUSION: The 3.7% rate of seroprevalence for CE in asymptomatic municipal waste collectors living in urban regions of Ahvaz suggests a high rate. The higher rate of infection among workers in district 5 is likely associated with the presence of stray and owned dogs in that area of the city. A prolonged exposure to contaminated soil, the lack of awareness about the risk of diseases that can be transmitted through waste and the lack of general availability of suitable personal protective equipment for waste collectors might cause infectious diseases.

Paleoparasitology in Iran: A Review.

Paleoparasitology was created to trace and recover the natural development of parasites as well as the origin of infectious diseases. Paleoparasitology is defined as the study of parasites in ancient material and their interactions with hosts and vectors. Advances in the field have helped to open up new prospects for anthropologists, archaeologists, biologists and medical scientists. In recent years, Iranian parasitologists and biologists have developed immense interest in this field. One of the first human settlements on earth was established in Iran and there is extensive evidence of early human life in this ancient land. Therefore, the aim of the review was to assess paleoparasitological research conducted in Iran in order to facilitate the discovery of the origin of infectious diseases in the region. English and Persian electronic databases including Web of Science, Sciencedirect, PubMed, Scopus, Google Scholar, Iran Doc, SID, Iran Medex and Magiran were employed as search engines (up to 2017) using the keywords: Iran, Islamic Republic of Iran, Parasitology, Parasites and Archaeology. According to the current review, the results of the parasitological study revealed the incidence of human and animal parasitic infection in Iran dating back to 8100 BC.

Genomic analysis of Blastocystis hominis isolates in patients with HIV-positive using locus SSU-rDNA.

Blastocystis hominis (B. hominis) is a protozoan zoonosis which clinical signs of infection with this parasite has been reported to be more severe in patients with weakened immune systems than healthy controls. So, the aim of the study was to evaluate genomic analysis of B. hominis isolates obtained from patients with HIV-positive using locus SSU-rDNA. At first, 268 stool samples were randomly collected from patients with HIV-positive referred to health centers of Khuzestan province, southwest of Iran. Formol-ether and direct smear techniques were used for the detection of parasitic agents. After extracting DNA, the samples were analyzed by the PCR method. Finally, the subtypes were determined by the sequencing and PCR methods. New samples were used for the preparation of positive control sample; they were cultured in coagulant-serum biphasic cultivation media. Of 268 stool samples, 33 (12.3%) cases were detected positive for B. hominis using Formol-Ether technique but 51 (19%) cases were positive using molecular method. The most common isolates were related to the subtype III with 29 positive cases (56.8%), then, genotype I with 11 (21.6%) cases, 6 cases (11.8%) with genotype II, 3 (5.9%) combined cases with genotypes I and III as well as 2 cases (3.9%) with genotype VI. There was a significant difference between two groups of HIV-positive patients (infected with the parasite and/or without the parasite) in the term of the mean of TCD4-positive cells. The results indicated a relatively high prevalence of B. hominis in HIV-positive patients as well as our findings may represent that the number reduction of TCD4-positive cells has an effective role in the increased risk of the parasitic infection in HIV-positive patients.

Molecular diagnosis of microsporidia strains in slaughtered cows of southwest of Iran.

Microsporidia is often considered as an opportunistic infection in patients with impaired immune systems such as patients with acquired immune deficiency syndrome, transplant recipients, children and old people. Due to the ability of the parasite to transmit from animals to human as well as the increasing prevalence of parasitic infections and immune deficiency diseases; therefore, the aim of this study was to evaluate molecular diagnosis of microsporidia strains in slaughtered cows of southwest of Iran. Initially, 256 stool samples of cows were collected from 5 regions of Khuzestan province, southwest of Iran and stained by modified trichrome (weber). Then, the parasite spores were examined by optical microscope. Total DNA was extracted from samples using DNA extraction kit for stool (Bioneer) and evaluated by multiplex/nested PCR method. The products of nested-PCR were explored by RFLP method using restriction enzyme MnlI. For genotyping, positive samples of RFLP were sequenced. Of 256, 21 and 48 samples were found positive by the staining and nested PCR tests, respectively. Of 48, 36 samples were Enterocytozoon bieneusi with genotype frequency D (22), J (9) and M (5). Also 9 were detected as Encephalitozoon species among which 2 were E. cuniculi, 6 were E. intestinalis and 1 was E. hellem Eventually, 3 samples were found positive for both Enterocytozoon and Encephalitozoon. The results showed that cows can be a source of microsporidia infections. Due to the zoonotic importance of this parasite and its ability to transmit from animals to humans; the detection and species determination of the parasite seems essential. The highest risk of infection is for individuals with impaired immune systems.

Prevalence of Cryptosporidium species isolated from HIV/AIDS patients in southwest of Iran.

This study aimed to determine the prevalence and species of Cryptosporidium among HIV/AIDS patients in southwest of Iran. Two hundred fifty faecal samples from HIV patients were examined for the presence of Cryptosporidium oocysts using a conventional coproscopic approach. Such oocysts were detected in 18 (7.2%) out of 250 faecal samples. Genomic DNAs from 250 samples were then subjected to a nested-PCR-RFLP technique targeting different loci of 18S rRNA gene for species identification. Out of 250 samples, 27 (10.8%) were positive for different Cryptosporidium spp; Restriction patterns resulting from the digestion of the nested amplicon with restriction endonucleases VspI and SspI showed that C. parvum (70.38%) was the most prevalent species, followed by C. hominis (25.92%) and C. meleagridis (3.7%), respectively. The mean CD4+ T-cell count was 215 cells/µL. There was a strong association between cryptosporidiosis and CD4+ T-cell count (P = 0.000) with the highest prevalence recorded among patients with CD4+ T-cell count < 200 cells/µL. This confirms that there is a low opportunity for this parasite to get established as the patients CD4+ T-cell count increases. Also HIV infection increased the risk of having Cryptosporidium. Our epidemiological findings are useful for any preventive intervention to control disease diffusion.