Epidemiological and Diagnostic Features of Blastocystis Infection in Symptomatic Patients in Izmir Province,Turkey

Background

The aims of this study were to identify Blastocystis subtypes (STs) in a cohort of Turkish patients with various gastrointestinal symptoms using a novel Real Time PCR method developed recently for Blastocystis detection and assess the relationship between Blastocystis STs and patient symptoms.

Methods

Totally, 617 stool samples of patients with gastrointestinal symptoms were examined with microscopy and inoculated in Jones medium. Blastocystis-positive samples were further assessed to identify coinfections with other possible pathogens, including bacteria and viruses. Diagnostic efficacies of microscopy, culture and Real-Time PCR were compared. PCR products were sequenced to identify the subtypes of Blastocystis isolates.

Results

Totally 94 (15.24%) samples were positive for Blastocystis after all methods. Among these, 83 of 94 (88.3%) samples were identified with all methods, while 11 were positive only with Real Time PCR. Diarrhea and abdominal pain were the leading symptoms in the patients. The only pathogenic agent identified in 76 of 94 (80.9%) patients was Blastocystis. Subtype 3 was the leading Blastocystis subtype (44.6%), while subtypes 6 and 7 were firstly isolated from symptomatic patients in our region.

Conclusion

Comparison of three diagnostic methods indicated Real Time PCR as the most sensitive and specific method. Blastocystis was the only pathogenic agent among symptomatic patients, with subtype 3 being predominant. Patients with subtypes 6 and 7 need further assessments concerning the zoonotic potential of Blastocystis.     

First Molecular Identification of Sarcocystis ovicanis (Protozoa,Apicomplexa) in the Brain of Sheep in Iran

Background

The objective of the present study was to survey the presence of Sarcocystis in sheep’s brain in North Khorasan Province.

Methods

In general, 80 samples of sheep’s brain were collected from slaughtered sheep in slaughterhouses of North Khorasan Province. Tissue digestion method was used for observing bradyzoites in tissues. Histopathological processing tracing Sarcocystis and ensuing structural change in the brain tissue were conducted. PCR analysis was conducted on all the brain samples. Sequencing was done for one PCR product. Genotype was identified by Blast search and homology analysis.

Result

Sarcocystis spp. was found in one of the brain samples (1.25%) using tissue digestion method. The presence of bradyzoite was also confirmed in the prepared histopathological sections. PCR analysis was positive in one of samples. Genotyping of one sample proved that Sarcocystis species was Sarcocystis ovicanis and the nucleotide sequence of this parasite was deposited in the GenBank database under accession number No.{"type":"entrez-nucleotide","attrs":{"text":"KF489431","term_id":"540198437","term_text":"KF489431"}}KF489431.

Conclusion

Sarcocystis ovicanis can involve brain tissue of sheep and consequently causes clinical symptoms.     

Prevalence of Balantidium coli Infection in Bred Rhesus Monkeys (Macaca mulatta) in Guangxi,southern China

Background

Balantidium coli infects humans, primates and pigs, causing serious diarrhea and dysentery. Little information on the prevalence of B. coli in primates is available in China. This investigation was conducted to determine the prevalence of B. coli infection in bred rhesus monkeys in Guangxi Zhuang Nationality Autonomous Region (GZNAR), southern China.

Methods

A total of 120 fecal samples were collected from rhesus monkeys bred in cages in GZNAR and B. coli cysts and/or trophozoites were examined microscopically after sedimentation with water in May 2013.

Results

(64.2%) samples were tested positive. The prevalence was 65% (39/60) and 63.3% (38/60) in female and male monkeys, respectively. 80% (48/60) cages in this nonhuman primate center were positive for B. coli.

Conclusion

The present survey revealed high circulation of B. coli in bred rhesus monkeys in GZNAR, which poses potential threats to animal and human health.     

Emerging Intestinal Microsporidia Infection in HIV+/AIDS Patients in Iran: Microscopic and Molecular Detection

Background

Species of Microsporidia have been known as opportunistic obligate intracellular parasites particularly in immunocompromised patients. Enterocytozoon bieneusi is one of most prevalent intestinal microsporida parasites in HIV⁺/AIDS patients. In this study, intestinal microsporidia infection was determined in HIV⁺/AIDS patients using microscopic and molecular methods.

Methods

Stool samples were collected from HIV⁺/AIDS patients during 12 months. All of the stool specimens washed with PBS (pH: 7.5). Slim slides were prepared from each sample and were examined using light microscope with 1000X magnification. DNA extraction carried out in microscopic positive samples. DNA amplification and genus/species identification also performed by Nested-PCR and sequencing techniques.

Results

From 81 stool samples, 25 were infected with microsporidia species and E. bieneusi were identified in all of positive samples. No Encephalitozoon spp. was identified in 81 collected samples using specific primers.

Conclusion

E. bieneusi is the most prevalent intestinal microsporidia in immunocompromised patients of Iran. On the other hand, Nested-PCR using specific primers for ssu rRNA gene is an appropriate molecular method for identification of E. bieneusi.     

Molecular and Parasitological Study of Cryptosporidium Isolates From Cattle in Ilam,West of Iran

Background

Cryptosporidiosis is one of the most important parasitic infections in human and animals. This study was designed for survey on the prevalence of Cryptosporidium infection in farms of Ilam, west of Iran, using parasitology method and genotyping by Nested PCR-RFLP.

Methods

Fecal samples of 217 cattle were collected fresh and directly from the rectum of cattle. All of the samples were examined by microscopic observation after staining with modified Ziehl-Neelsen (MZN). Genomic DNA extracted by using EURx DNA kit. A Nested PCR-RFLP protocol amplifying 825 bp fragment of 18s rRNA gene conducted to differentiate species and genotyping of the isolates using SspI and VspI as restriction enzymes.

Results

The prevalence of Cryptosporidium infection in cattle using both methods is 3.68%. Most of the positive cattle were calves under six months. Species diagnosis carried out by digesting the secondary PCR product with SspI that C. parvum generated 3 visible bands of 448, 247 and 106 bp and digested by VspI restriction enzyme generated 2 visible bands of 628 and 104bp. In this investigation all of the positive samples were Cryptosporidium parvum.

Conclusion

C. parvum (bovine genotype) detected in all positive cattle samples in Ilam, west of Iran. The results of the present study can help for public health care systems to prevention and management of cryptosporidiosis in cattle and the assessment of cattle cryptosporidiosis as a reservoir for the human infection.     

Isolation of Small Number of Cryptosporidium parvum Oocyst Using Immunochromatography

Background

Cryptosporidium parvum causes severe gastroenteritis in immunocompromised human and new borne animals. The organism can be transmitted through water. Since small number of C. parvum is infectious, the aim of the present study was to develop a chromatography method for the isolation of C. parvum oocyst in samples with limited number of oocysts.

Methods

Antibody was prepared against whole antigen from C. parvum oocysts, the achieved Ab bound to the sepharose 4B and used for the isolation of oocysts. Antibody against P23 bound to the sepharose 4B, used also for the isolation of C. parvum oocyst. In comparison to these both methods, 2 traditional methods (Salt floatation and 55% sucrose floatation) were also performed.

Results

Both chromatography methods could bind oocysts with capacity depends on the column size. The isolated oocysts were free of bacteria. Our results showed that the traditional methods are useful for the isolation of oocysts from feces, in its smear stained with ziehl-nelsen, at least 3 oocyts are detectable in each microscopic field under 1000 X magnification. In contrast to the chromatography methods, the bacterial contamination was always observed in oocysts isolated with traditional methods.

Conclusion

Immunochromatography could be used for the successful isolation of C. parvum oocysts from the samples containing limited number of oocysts.     

A Multi-Locus Study of Cryptosporidium Parasites Isolated From Patients Living In Iran,Malawi, Nigeria,the United Kingdom,and Vietnam

Background

Cryptosporidium species are important cause of diarrheal diseases in both developing and developed countries. This study aimed to compare the performance of several molecular methods for identification of Cryptosporidium species, and to detect genetic variation among each of these species isolated from Iran, Malawi, Nigeria, Vietnam and the United Kingdom.

Methods

The oocysts DNA samples were derived from 106 Cryptosporidium positive feces. Polymerase chain reaction, PCR- restriction fragment length polymorphism and DNA sequence analysis of the 18S rRNA and the Cryptosporidium oocysts wall protein genes; PCR and DNA sequence analysis of a fragment of 70 kDa heat shock protein and 60 kDa glycoprotein genes were carried out.

Results

Based on these analysis, three species of Cryptosporidium including C. hominis, C. parvum and C. meleagridis, and both C. hominis and C. parvum were found in Iranian and the UK samples, respectively. Also, three C. hominis (Ib, Ib3& Id) and three C. parvum (IIa, IIc & IId) subtypes were identified by sequence analysis of the GP60 gene. Of these, C. hominis Ib was predominant and interestingly, one subgenotype (C. hominis Ib A10G2) accounted for the majority of the samples.

Conclusion

The current study demonstrates the complex subtypes of Cryptosporidium isolates in both developing and developed countries. This is the first report of C. parvum IId subgenotype and three new subtypes of C. parvum IIa in the UK, a new subtype of C. hominis Id from Malawi; and the first multi-locus study of three species of Cryptosporidium in human from Iran.     

Isolation and Identification of Naegleria Species from Environmental Water in Changchun,Northeastern China

Background

Naegleria is a free-living amoeba, and pathogenic Naegleria may pose a health risk to people exposed to recreational water. Our objective in this study was to determine if there are pathogenic amoebae in environmental water samples from Changchun, Northeastern China.

Methods

During July to September 2012, a total of 70 water samples were collected from Changchun, Northeastern China, and Naegleria was enriched by in vitro culture and detected by PCR using Naegleria genus-specific primers. Resulting PCR products were sequenced and phylogenetically analyzed to identify Naegleria species.

Results

Naegleria was detected in 65 (92.9%) of 70 water samples. DNA sequence and phylogenetic analyses based on the internal transcribed spacer (ITS) rDNA sequences revealed four Naegleria species, including N. pagei (n = 24) and N. Australiensis (n = 18), N. clarki (n = 13) and N. gruberi (n = 10), in which N. australiensis is pathogenic to mice. But the pathogenic species N. fowleri was not detected.

Conclusion

This is the first report on Naegleria species in Northeastern China, showing that almost all environmental water samples were contaminated with Naegleria, including N. pagei, N. Australiensis, N. clarki and N. gruberi, which should be considered a potential public health threat.     

Seroprevalence of Toxoplasma gondii Infections in Pregnant Women in Gorgan City,Golestan Province,Northern Iran-2012

Background

Toxoplasma gondii is one of the most prevalent parasites of human and warm- blooded animals. Toxoplasmosis is important especially in two groups: pregnant women and immunocompromised patients. If women acquire the primary infection during the pregnancy, it would be life threatening or remains severe disorders for the fetus. This study was performed to evaluate the seroprevalence of T. gondii infection in pregnant women referred to Health Center in Gorgan City, Golestan Province, northern Iran.

Methods

Serum samples were collected from pregnant women referred to Health Center in Gorgan City, south eastern Caspian Sea. Anti- Toxoplasma IgG and IgM antibodies were determined by commercially ELISA kits and the relation of infection with socio-demographic and risk factors such as age, education, occupation, cat ownership, soil contact and some other factors was studied.

Results

From 555 tested sera of pregnant women referred to Health Center in Gorgan, 39.8% had IgG antibodies against T. gondii and 3.4% were positive for IgM antibodies. A significant correlation was seen between T. gondii infection with age and soil contact.

Conclusion

About 60% of pregnant women in Gorgan City are seronegative against T. gondii, so they should considered as at risk persons.     

Comparison of Three Staining Methods for the Detection of Intestinal Microspora Spp

Background

This study aimed to compare three staining methods including: Calcofluor white, Chromotrope and Quick Hot Gram chromotrope used in diagnosis of intestinal microsporidial spores.

Methods

One hundred and seventy five stool specimens were collected from patients referred to Laboratory of Intestinal Protozoology at the School of Public Health, Tehran University of Medical Sciences during 2012-2013. All of specimens were evaluated by nested PCR. The formalin–fixed stool samples were prepared from each specimen and dried at room temperature for 10 min, followed by 10 min methanol fixation. All the collected stool samples were evaluated blindly by calcofluor white, Chromotrope and Quick Hot Gram chromotrope staining methods separately.

Results

Microsporidial spores were recognized using Chromotrope, Quick Hot Gram chromotrope and Calcofluor white, in16 of 18 (88.8%), 17 of 18 (94.4%) and 18 of 18(100%) samples that were positive by nested PCR respectively. Regarding 14 stool samples that were negative by nested PCR, 14 cases were negative by chromotrope and Quick hot Gram chromotrope and 13 samples were negative by Calcofluor white. One discordant sample interpreted as false positive.

Conclusion

Calcofluor white staining had the best performance for the detection of intestinal Microsprora spores and can be used as initial screen test for the detection of intestinal Microspora spp.     

Genetic Identification of Trichomonas vaginalis by Using the Actin Gene and Molecular Based Methods

Background

Trichomonas vaginalis is the agent of urogenital tract infection that causes human trichomoniasis with some serious health complications. More understanding about genetic features of the parasite can be helpful in the study of the pathogenesis, drug susceptibility and epidemiology of the infection. For this end, we conducted analysis of the actin gene of T. vaginalis by applying the PCR-SSCP (PCR-Single Stranded Conformational Polymorphism) and nucleotide sequencing method.

Methods

Fifty T. vaginalis samples were collected from 950 women attending gynecology clinics in two cities of Iran, Hamadan and Tehran, from November 2010 to July 2011. After axenisation of isolates, all samples subjected to PCR-SSCP and nucleotide sequencing.

Results

According to the SSCP banding patterns and nucleotide sequencing, seven sequence types were detected among the isolates. Alignment of the nucleotide sequences showed five polymorphic sites in the different strain types. Amino acid substitution was not observed in the nucleotide sequence translation of the all sequences.

Conclusion

The actin gene analysis represents genetic diversity of T. vaginalis and it suggests that various strains can be responsible for clinically different trichomoniasis in infected individuals. It is expected that further studies will be conducted to increase our knowledge about relationship between the actin gene polymorphism and different biological behavior of the parasite.     

Incidence of Giardia lamblia Subspecies by PCR-RFLP in Stool Specimens of Hospitalized Children at Urmia Mutahhari Hospital,West Azerbaijan Province,Iran

Background

Giardia lamblia is one of the most prevalent intestinal flagellate protozoa that infects a wide range of vertebrate hosts causing severe intestinal disorder in children.This study was performed to determine subspecies of G.lamblia by the PCR-RFLP method, targeting the glutamate dehydrogenase(gdh)locus, in hospitalized children at Urmia Mutahhari Hospital, West Azerbaijan Province,Iran and determining the infection transformational storages in this area.

Methods

Overall, 720 stool specimens were collected from the hospitalized children, 34 samples were positive and Giardia cysts were detected under the microscope. Cysts were partially purified by the sucrose density gradient method and then washed with sterile distilled water to remove effectively the PCR inhibitors. Genomic DNA of G. lamblia isolates was extracted by freeze-thaw cycles followed by phenol/ chloroform/isoamyl alcohol method. The single step PCR-RFLP assay was used to differentiate the assemblages between A and B, which were found in humans. In this method, 432 bp expected size was amplified, and then for detection of subspecies, specific restriction RsaI and BspLI enzymes were used.

Results

Totally 34 samples were positive in terms of Giardia cyst out of 720 examined samples microscopically, so the parasite spread rate is reported 4.72%. Analysis PCR-RFLP on these samples revealed that 28 samples (93.3%) have the genotype BIII and 2 samples (6.7%) belong to the subgroup BIV.

Conclusion

PCR-RFLP is a proper analytical method for determining the genotype among parasite types, using the glutamate dehydrogenizes zone’s genes. Based on the results, an animal origin of infection cycle is suggested.     

Acanthamoeba species in Swimming Pools of Cairo,Egypt

Background

The free-living amoebae Acanthamoeba spp. have been recognized as etiologic agents of amoebic encephalitis, keratitis, otitis, lung lesions and other skin infections mainly in immuno-compromised individuals. The purpose of this study is to detect the presence of Acanthamoeba in swimming pools in Egypt using a polymerase chain reaction (PCR) method.

Methods

Water samples were collected from 10 different swimming pools in Cairo, Egypt. Samples were cultured on non-nutrient agar for the detection of Acanthamoeba isolates that were confirmed by PCR amplification using genus specific primers. The molecularly confirmed Acanthamoeba isolates were morphologically identified to the species level.

Results

Members of genus Acanthamoeba were detected in 49.2% of the examined swimming-pool water samples. Morphologically, six Acanthamoeba species were isolated from the examined swimming pool water namely A. polyphaga, A.castellanii, A. rhysodes, A. mauritaniensis, A. royreba and A. triangularis. All the identified species of Acanthamoeba were molecularly confirmed to be related to the genus Acanthamoeba.

Conclusion

The isolated species of Acanthamoeba could provoke variable degrees of infections to the swimmers. The culture method is cheaper and easier than PCR techniques that are faster for the detection of free-living amoebae     

Molecular Survey of Toxoplasma gondii in Sheep,Cattle and Meat Products in Chaharmahal va Bakhtiari Province,Southwest of Iran

Background

Toxoplasmosis is a worldwide spread disease. The present study examined the prevalence of Toxoplasma gondii infection among animals of edible meat (cattle and sheep) in Chaharmahal va Bakhtiari Province (Southwest of Iran) in 2012. Furthermore, we attempted for the first time to identify this parasite from the meat products in the province.

Methods

The tongue, brain, femur muscle and liver of 50 sheep and 70 cattle as well as 50 samples of meat products were selected and collected to perform molecular survey using Nested-PCR method.

Results

Of the studied sheep, 38% were infected. The infection rate in the age groups under 1 year, 1-2 years, and more than 2 years was 25%, 35.29% and 52.94%, respectively. The infection rate in femur muscle, brain, liver and tongue was 28%, 32%, 30% and 16%, respectively. Of the studied cattle, 8.57% were infected. The infection rate in the age groups 1-2 years, 2-4 years, and more than 4 years was 3.7%, 9.09% and 14.28%, respectively. Sheep was infected 6 times more than cattle (OR = 6.53 CI = 2.374-18.005).The infection rate among samples of meat products was 12% (6 samples out of 50 samples).

Conclusion

Due to the high rate of this parasitic infection among the slaughtered animals as well as meat products in this region, the use of infected material can be one of the main risk factors of transmission of the parasite to humans.     

Integrating Morphology,Breeding Ground and Mitochondrial COI Gene Analysis for Species Identification of Bellamya lithophaga (Gastropoda: Viviparidae) in China

Background

Angiostrongylus cantonensis is a zoonotic public health concern that causes human severe eosinophilic meningitis in Southeast Asia and China. As a medically important intermediate host of A. cantonensis, Bellamya lithophaga (Gastropoda: Viviparidae) is often confused with other morphologically similar sibling species of genus Bellamya, such as B. aeruginosa and B. purificata in the past. Hence, the aim of the present study was to investigate evidences to discriminate these equivocal Bellamya species.

Methods

This study was carried out by getting Bellamya snail samples from Fujian Province in the South-East of China. The snail morphological features, breeding grounds and phylogenetic relationship according to mitochondrial cytochrome c oxidase subunit I (COI) gene marker were analyzed.

Results

Based on external morphology, radular shape and cusp formula, as well as major breeding environment, B. lithophaga could be distinguished from B. aeruginosa, B. purificata. The phylogenetic tree also unconfirmed that B. lithophaga belongs to a different genetic clade from other morphologically similar species.

Conclusion

Our findings demonstrate the significant differences in B. lithophaga and other sibling species, which supports the traditional species delimitation in the genus Bellamya.     

The Rate of Plasmodium vivax Infectivity within Gloucose-6-Phosphate Dehydrogenase (G6PD) Deficient Individuals in Hormozgan Province,Iran

Background

One of the most important enzymatic disorders that interact with malaria is deficiency of G6PD (Gloucose-6-phosphate dehydrogenase). This enzyme protects red blood cells from hydrogen peroxide and other oxidative damages. Distribution of this enzyme deficiency usually accompanies with low level distribution of malaria disease in most malarious areas. So this hypothesis may be considered that the G6PD deficiency could be protective against malaria.

Methods

Totally 160 samples were taken from vivax malaria infected and non-infected individuals. Preparing blood smears and quantitative test for G6PD deficiency were employed for all of the samples. To ensure accuracy of the malaria in negative samples besides using microscopical examination, semi-nested multiplex PCR was also performed for the two groups.

Results

In microscopical examination 36 and 124 samples were vivax malaria positive and negative respectively. Out of 36 P.vivax positive cases 3 (8.3%) cases were detected to be G6PD deficient versus 30 (24.2%) cases out of 124 P. vivax negative cases. The results showed a significant differentiation between P. vivax positive and P. vivax negative cases in the rate of G6PD deficiency (3/36 in positive cases versus 30/124 in negative cases) (P<0.05).

Conclusion

vivax malaria positive individuals with G6PD deficiency showed too mild symptoms of Malaria or even asymptomatic.     

Canine Babesiosis in China Caused by Babesia gibsoni: A Molecular Approach

Background

To provide a point of reference to study the epidemiology and clinical expression of canine babesiosis in China.

Methods

A total of 30 dogs infected with canine babesiosis were evaluated by mean of clinical history, physical examination, hematological, restriction fragment length polymorphism of PCR products (PCR-RFLP) and sequencing analysis.

Result

The most prevalent clinical abnormalities were lethargy (100%), anorexia (100%), pale or icteric mucous membranes (80%), fever (70%) and dark urine (70%). Hematology parameters revealed that anemia and thrombocytopenia were the major abnormalities in blood of dogs infected with canine babesia. The results of PCR-RFLP and sequencing analysis indicated that B. gibsoni was the main species responsible for canine babesiosis cases at the time of the study in Nanjing, China.

Conclusions

The results provide valuable information for better understanding of the epidemiology of canine babesiosis in China.     

Molecular Identification of Giardia duodenalis Isolates from Fars Province,Iran

Background

Giardia duodenalis is one of the most common human intestinal protozoan parasites worldwide and is endemic throughout the world with a vast range of mammalian hosts. The present study aimed to identify the prevalence of G. duodenalis isolates and determine the most common of its assemblages in the patients referring to health centers and hospitals in Fars province, Iran that will be subjected to further molecular investigation.

Methods

We collected 1000 human fecal samples from health centers and hospitals in Shiraz, Iran in a one year period from September 2009 to August 2010. Microscopic examination for the presence of G. duodenalis cysts and trophozoites was performed by direct wet mount before and after the concentration techniques. Extraction of DNA was performed by Phenol-Chloroform-Isoamylalcohol (PCI). G. duodenalis-positive specimens were analyzed by PCR. A fragment of the SSU-rDNA (292 bp) gene was amplified by PCR using the forward primer RH11 and the reverse primer RH4. Genotyping was performed using sequence analysis of G. duodenalis glutamate dehydrogenase gene using primers GDHeF, GDHiF, and GDHiR.

Results

The prevalence of Giardia infection was 10.7% (107/1000) examined based on microscopic examination. PCR identified 80% (40/50) of the samples as positive for G. duodenalis based on SSU-rDNA amplification on sucrose gradient samples. Besides, genotyping results indicated 32 isolates (80%) as assemblage AII and 8 isolates (20%) as assemblage BIII and BIV based on the DNA sequence analysis of the glutamate dehydrogenase locus of G. duodenalis.

Conclusion

The findings of this study emphasize that Iran (Fars Province) is a favorable area for giardiasis with an anthroponotic infection route.     

Molecular Detection of Malaria in South Punjab with Higher Proportion of Mixed Infections

Background

Malaria is well known for its fatalities worldwide, Plasmodium vivax and the Plasmodium falciparum are the two important species of malaria reported from Pakistan and creating lots of morbidities across the country.

Method

Study was conducted to determine the Surveillance of malaria in South Punjab by microscopy and Polymerase chain reaction (PCR).

Result

samples out of 100 patients were found positive for malarial parasites. One patient was found with mixed infection, whereas P. falciparum and P. vivax infections were detected in 17 and 22 patients, respectively. In nested PCR, genus-specific primers for Plasmodium species. in round 1 and species-specific primers for P. falciparum and P. vivax in round 2 were used. By the application of PCR 41% were found to be infected by Plasmodium spp. Among Plasmodium positive patients: mixed, P. falciparum and P. vivax infection were detected in 10, 15 and 16 patients, respectively. Thirty nine microscopically positive patients confirmed to have Plasmodium spp. One negative by PCR, 2 microscopically negative patients had shown Plasmodium spp. infection (P. falciparum and P. vivax) by PCR. In total samples, P. falciparum, P. vivax and mixed infection accounted for 36.6%, 39.0% and 24.3%, respectively.

Conclusion

Microscopy was found deficient for interpretation of mixed infections, low parasitaemia, and species specific diagnosis. The sensitivity, specificity and efficacy of nested PCR was calculated 95%, 98% and 97%, respectively, showing PCR as a more effective and efficient diagnostic tool for malaria.     

Molecular and Serological Detection of Acute and Latent Toxoplasmosis Using Real-Time PCR and ELISA Techniques in Blood Donors of Rafsanjan City,Iran, 2013

Background

The differentiation between acute and latent forms of the Toxoplasma gondii (T. gondii) infection is still considered as a complicated issue. This study was aimed to elucidate the status of infection in the blood donors and the probable importance of blood transfusion in the transmission of the infection through detecting both immunological and genetic markers of acute and latent infection.

Methods

Totally 235 blood samples from blood donors were collected. The levels of anti-T. gondii IgG and IgM antibodies were examined by specific ELISA kits. cDNA were synthesized from total extracted mRNA molecules from the serum samples and SAG1 gene, specific for tachyzoite form, were amplified using Real-Time PCR technique. Demographic information of study subjects including their gender, age, job, and habitat were recorded.

Results

Out of 235 serum samples, 80 (34.04%) and 4 (1.71%) were positive regarding anti-T. gondii IgG and IgM antibodies, respectively. Real-Time PCR results showed that 14 out of 200 (6.97%) of blood donor had mRNA molecules of SAG1 gene. The positive results of Real-Time PCR of SAG1 in female gender and housekeepers were significantly higher than those of male gender and other job categories.

Conclusion

The prevalence of chronic and acute infection is high in Iranian blood donors. Additionally, evaluation of antibodies could not be reliable, because several donors negative for anti-T. gondii IgM antibodies had detectable SAG1 mRNA molecules. Hence, it seems that molecular diagnostic tests are essential to detect acute infections.