Drug Resistance Pattern of Mycobacterium tuberculosis Isolates From Patients Referred to TB Reference Laboratory in Ahvaz

Objectives

Tuberculosis remains one of the top three infectious disease killers. The prevalence of multidrug-resistant tuberculosis (MDR-TB) has increased substantially in the past 20 years. When drug resistance is not detected, MDR-TB patients cannot access life-saving treatment; this puts their communities at risk of ongoing MDR-TB transmission. We aimed to determine the patterns of resistance to antituberculosis drugs among Mycobacterium tuberculosis isolates from Khuzestan province in Iran.

Methods

A total of 850 clinical specimens from patients suspected of active TB were cultured in 2015. Drug susceptibility testing to the first line antiTB drugs for culture positive MTB was performed on Lowenstein–Jensen medium using the proportion method.

Results

Of 850 cultured specimens, 272 (32%) were culture positive for mycobacteria. Of 64 MTB isolates that were analyzed by the proportion method, 62 (96.8%) were pan-susceptible and two (3.1%) were MDR.

Conclusion

An important way to prevent the emergence of MDR and XDR TB, and the principles of full implementation of the strategy is directly observed treatment, short-course (DOTS). The efficient diagnosis and timely treatment of MDR-TB patients can prevent disease transmission, reduce the risk of drug resistance developing, and avoid further lung damage.     

Boosting with a DNA vaccine expressing ESAT-6 (DNAE6) obliterates the protection imparted by recombinant BCG (rBCGE6) against aerosol Mycobacterium tuberculosis infection in guinea pigs

Owing to its highly immunodominant nature and ability to induce long-lived memory immunity, ESAT-6, a prominent antigen of Mycobacterium tuberculosis, has been employed in several approaches to develop tuberculosis vaccines. Here, for the first time, we combined ESAT-6 based recombinant BCG (rBCG) and DNA vaccine (DNAE6) in a prime boost approach. Interestingly, in spite of inducing an enhanced antigen specific IFN-γ response in mice, a DNAE6 booster completely obliterated the protection imparted by rBCG against tuberculosis in guinea pigs. Analysis of immunopathology and cytokine responses suggests involvement of an exaggerated immunity behind the lack of protection imparted by this regimen.     

Comparison of antibody and cell-mediated immune responses of foals and adult horses after vaccination with live Mycobacterium bovis BCG

Background

Equine neonates have reduced humoral and cell-mediated immune responses compared to adult horses after administration of killed vaccines. As a basis for this study, we hypothesized that newborn foals can mount strong immune responses after vaccination with live Mycobacterium bovis BCG.

Methods

Healthy 4-day-old foals (n = 7), 4-month-old foals (n = 7) and adult horses (n = 6) were vaccinated once with live M. bovis BCG. Age-matched animals (n = 5 per group) were used as unvaccinated controls. Relative vaccine-specific immunoglobulin concentrations and whole blood mRNA expression of IFN-γ, IL-4, and IL-10 were measured prior to and 2, 4, 6, and 8 weeks after vaccination. Eight weeks after vaccination, delayed type hypersensitivity (DTH) responses were assessed by measuring the increase in double skin thickness after intradermal injection of purified protein derivative.

Results

Both groups of foals and adult horses responded with a significant increase in vaccine-specific total IgG, IgGa, IgGc, IgG(T), and IgM concentrations. In contrast, only adult horses mounted significant IgGb responses. Vaccine-specific concentrations of total IgG and IgGa were significantly higher in adult horses than in 4-day-old foals whereas IgGc responses were significantly higher in 4-day-old foals than in the other two age groups. Adult horses had significantly higher basal IFN-γ and IL-4 mRNA expression than both groups of foals but vaccination with M. bovis BCG did not significantly increase expression of these cytokines, regardless of age group. Immunized horses had significantly higher DTH responses than age-matched unvaccinated controls. DTH responses were significantly greater in both groups of vaccinated foals than in vaccinated adult horses.

Conclusion

Despite a naïve immune system, newborn foals have the ability to mount robust antibody and cell-mediated immune responses to M. bovis BCG.     

Safety and reactogenicity of BCG revaccination with isoniazid pretreatment in TST positive adults

Rationale

Global tuberculosis (TB) control may require mass vaccination with a new TB vaccine, such as a recombinant bacille Calmette Guerin (BCG) or attenuated Mycobacterium tuberculosis (MTB). The safety profile of live mycobacterial vaccines in latently infected adults with prior infant BCG vaccination is unknown.

Objectives

Evaluate safety and reactogenicity of BCG revaccination, with or without isoniazid (INH) pretreatment, in adults with latent MTB infection (LTBI).

Methods

Eighty-two healthy, HIV uninfected, South African adults, with a BCG scar and tuberculin skin test (TST) diameter ≥15 mm, were randomized to receive 6 months of INH, starting either before, or 6 months after, intradermal revaccination with BCG Vaccine SSI (Statens Serum Institut, Copenhagen). Safety and reactogenicity data are reported through 3 months post BCG revaccination.

Results

Baseline characteristics were similar between treatment arms. Mean baseline TST diameter was 20 ± 4 mm. Seventy-two subjects received BCG revaccination. Injection site erythema (68%) and induration (86%) peaked 1 week after revaccination. Ulceration (76%) peaked at 2 weeks, and resolved by 3 months in all but 3 subjects. Diameter of ulceration was >10 mm in only 8%, but a residual scar was common (85%). No regional lymphadenitis or serious morbidity related to BCG was seen. Reactogenicity was not affected by INH pretreatment.

Conclusion

BCG revaccination of MTB infected adults is safe, well tolerated, and reactogenicity is similar to that of primary BCG vaccination. Clinical trials of live recombinant BCG or attenuated MTB vaccines may be considered in latently infected adults, with or without INH pretreatment (ClinicalTrials.gov identifier: NCT01119521).     

Molecular Typing of Mycobacterium intracellulare Using Pulsed-Field Gel Electrophoresis,Variable-Number Tandem-Repeat Analysis,Mycobacteria Interspersed Repetitive-Unit-Variable-Number Tandem Repeat Typing,and Multilocus Sequence Typing: Molecular Characterization and Comparison of Each Typing Methods

Objectives

Mycobacterium intracellulare is the major causative agent of nontuberculous mycobacteria-related pulmonary infections. The strain typing of M. intracellulare is important for the treatment and control of its infections. We compared the discrimination capacity and effective value of four different molecular typing methods.

Methods

Antibiotic susceptibility testing, hsp65 and rpoB sequencing, pulsed-field gel electrophoresis (PFGE), multilocus sequence typing (MLST), mycobacteria interspersed repetitive-unit-variable-number tandem-repeat analysis (MIRU-VNTR), and VNTR assay targeting 44 M. intracellulare isolates obtained from patients with pulmonary infections were performed.

Results

All the antibiotic susceptibility patterns had no association with the molecular and sequence types tested in this study; however, the molecular and sequence types were related with each other. PFGE gave best results for discriminatory capacity, followed by VNTR, MLST, and MIRU-VNTR.

Conclusion

The high discriminatory power of PFGE, VNTR, and MLST is enough for differentiating between reinfection and relapse, as well as for other molecular epidemiological usages. The MLST could be regarded as a representative classification method, because it showed the clearest relation with the sequence types.     

Effect of dietary protein and zinc on anti-mycobacterial antibody responses in guinea pigs

An enzyme immunoassay was developed to quantify the levels of antibodies reactive with purified protein derivative (PPD) at 3, 4, 5 and 6 wks following vaccination with Mycobacterium bovis BCG and initiation of diets deficient in either protein (LP) or zinc (LZ). Significant antibody levels were detected in all groups as early as 3 wks post-vaccination compared to non-vaccinated animals. Both LP and LZ guinea pigs had higher levels of antibody than animals consuming an adequate diet at 3 and 4 wks, respectively. By 6 wks post-vaccination, animals on the low protein diet had the highest antibody titers, followed by control, chow and LZ guinea pigs in order of decreasing titers. A highly significant positive correlation was detected between the levels of anti-mycobacterial antibodies and the number of viable BCG recovered from the lymph nodes draining the vaccination site. A significant inverse correlation was seen between antibody levels and PPD skin reactions. High-titer sera from protein-deficient, but not zinc-deficient, BCG-vaccinated pigs significantly suppressed the tuberculin reactions when passively transferred into normal, PPD-positive recipients.     

Evaluation of Dot Immunogold Filtration Assay (DIGFA) By Recombinant Protein EPC1 for Anti- Echinococcus granulosus IgG Antibody

Background:

Cystic echinococcosis is an important zoonosis caused by the Echinococcus granulosus with a substantial impact in human and animal health in endemic areas. The purpose of the present study was serodiagnosis optimizing of dog echinococcosis in order to achieve a rapid diagnostic method.

Methods:

Eight dogs were challenged with protoscoleces in order to have positive echinococcosis serum and 2 two-month old puppies were used as uninfected controls. Colloidal gold was prepared by controlled reduction of a boiling solution of chloroauric acid (H [AuCl4]) with sodium citrate and labeled with recombinant EPC1. Dot immunogold filtration assay (DIGFA) was developed by coating rEPC1 labeled colloidal gold on nitrocellulose membrane. The canine sera, taken three times including, 15, 28 and 35 days post infection were tested. A total of 30 serum samples including 24 sera from 8 infected dogs and 6 sera from 2 puppies were comparatively detected with both DIGFA and ELISA.

Results:

Gold labeled antigen, showed a dark purple dot with agglutination particles in positive sera and light purple dot without agglutination in negative sera. Among 30 serum samples, 23 were positive and 7 were negative with DIGFA and 24 were positive, 6 were negative with ELISA.

Conclusion:

DIGFA as a rapid and simple procedure could be utilized in quickly diagnosis of echinococcosis.     

Effect of Hominis Placenta on cutaneous wound healing in normal and diabetic mice

BACKGROUND/OBJECTIVES

The number of diabetic patients has recently shown a rapid increase, and delayed wound healing is a major clinical complication in diabetes. In this study, the wound healing effect of Hominis placenta (HP) treatment was investigated in normal and streptozotocin-induced diabetic mice.

MATERIALS/METHODS

Four full thickness wounds were created using a 4 mm biopsy punch on the dorsum. HP was injected subcutaneously at the middle region of the upper and lower wounds. Wounds were digitally photographed and wound size was measured every other day until the 14th day. Wound closure rate was analyzed using CANVAS 7SE software. Wound tissues were collected on days 2, 6, and 14 after wounding for H/E, immunohistochemistry for FGF2, and Masson''s trichrome staining for collagen study.

RESULTS

Significantly faster wound closure rates were observed in the HP treated group than in normal and diabetes control mice on days 6 and 8. Treatment with HP resulted in reduced localization of inflammatory cells in wounded skin at day 6 in normal mice and at day 14 in diabetic mice (P < 0.01). Expression of fibroblast growth factor (FGF) 2 showed a significant increase in the HP treated group on day 14 in both normal (P < 0.01) and diabetic mice (P < 0.05). In addition, HP treated groups showed a thicker collagen layer than no treatment groups, which was remarkable on the last day, day 14, in both normal and diabetic mice.

CONCLUSIONS

Taken together, HP treatment has a beneficial effect on acceleration of cutaneous wound healing via regulation of the entire wound healing process, including inflammation, proliferation, and remodeling.     

Immunotherapy Using Autoclaved L. major Antigens and M. vaccae with Meglumine Antimoniate,for the Treatment of Experimental Canine Visceral Leishmaniasis

Background

To evaluate immunotherapy against canine visceral leishmaniasis, Leishmania major antigen and heat-killed Mycobacterium vaccae (SRL172) were used as stimulators of immune defense mechanisms and the results were compared with standard chemotherapy meglumine antimoniate.

Methods

Nineteen mongrel dogs aging 1-3 years old were used in this experiment. Infection was carried out in 15 out of 19 dogs using L. infantum, isolated from a naturally infected poly-symptomatic dog.

Results

All the cases showed positive serologic results by direct agglutination test during 30-60 days following inoculation. In the first group, which was under chemotherapy (Glucantime^R), one of the members showed recurrence of the disease despite rapid effect of the therapeutic protocol. Immunotherapy using SRL172 caused complete cleaning of the parasite in group 2, but the speed was less than Glucantime. Immunotherapy using L. major antigen combined with M. vaccae in group 3 and combine administration of immunotherapy and chemotherapy in group 4 both were with relapsing of one case in each group. Group 5 and 6 were consisted of positive and negative control dogs, respectively.

Conclusion

Immunotherapy seems to be an adjuvant in treatment of canine leishmaniasis but it needs more investigation for final confirmation.     

Mathematical Modeling of Vibrio vulnificus Infection in Korea and the Influence of Global Warming

Objectives

To investigate the possible link between Vibrio vulnificus population size in seawater and water temperature.

Methods

We collected incidence and water temperature data in coastal regions of Korea and constructed a mathematical model that consisted of three classes; susceptible fish, infected fish available to humans, and infected humans.

Results

We developed a mathematical model to connect V. vulnificus incidence with water temperature using estimated bacterial population sizes and actual coastal water temperatures.

Conclusion

Increased V. vulnificus population sizes in marine environments may increase the risk of infection in people who eat at coastal restaurants in Korea. Furthermore, we estimated the near-future number of infected patients using our model, which will help to establish a public-health policy to reduce the disease burden.     

Sustained protection against tuberculosis conferred to a wildlife host by single dose oral vaccination

Background

Vaccination of wildlife against bovine tuberculosis (TB) is being considered by several countries to reduce the transmission of Mycobacterium bovis infection to livestock. In New Zealand, where introduced brushtail possums (Trichosurus vulpecula) are the major wildlife hosts, we have previously shown that repeat applications of a lipid-encapsulated oral bacille Calmette-Guerin (BCG) vaccine reduce the incidence of naturally acquired TB in wild possums. Here we extend this conceptual demonstration to an operational level, assessing long-term protection against TB conferred to free-living possums by a single oral immunisation.

Methods

Possums in a non-TB area were randomly allocated to receive lipid-formulated BCG vaccine or remained unvaccinated. After initial trials to assess vaccine immunogenicity and establishment of protection within the first year post-vaccination, 13 individuals of each treatment group were relocated to a biosecurity facility and challenged (at 28 months post-vaccination) by subcutaneous injection of virulent M. bovis.

Results

Vaccine immunogenicity and short-term protection were confirmed at 2 months and 12 months post-vaccination, respectively. In the long-term assessment, vaccinated possums had significantly reduced bacterial counts in peripheral lymph nodes compared to controls, with 0.6–2.3 log₁₀-fold reductions in M. bovis burdens.

Discussion

The magnitude of protective response by possums to experimental challenge at 28 months post-vaccination is known to equate to a high degree of protection against natural infection in this species. With techniques for oral bait delivery well advanced, the longevity of protection demonstrated here shows that an operable wildlife vaccine against TB is feasible.     

Characterization and immune effect of the hepatitis B–BCG combined vaccine for using a needle innoculation

Objective

To prepare the hepatitis B–Mycobacterium bovis Bacillus Calmette-Guérin combined vaccine (HB–BCG combined vaccine) and resolve a needle problem of the two kinds of hepatitis B vaccine (HB vaccine) and M. bovis Bacillus Calmette-Guérin (BCG) for the innoculation.

Methods

The hepatitis B surface antigen (HBsAg) was prepared by the genetic engineering technique, BCG was produced using routine biological technique, and then the finished products of the HB–BCG combined vaccine were processed on the above foundation. The content of HBsAg was measured by Enzyme linked immunosorbent assay (ELISA), the immune effect of BCG was detected by purified protein derivative (PPD) test. Cellular immune response, safety, partial poison and allergy were tested. The stability of HB–BCG combined vaccine was detected by ELISA and viable count method.

Results

The two kinds of antigens (HBsAg and BCG) had good compatibility. The comparison on immune effects of HB–BCG combined vaccine and BCG showed no significant difference. The comparison on immune effects of HB–BCG combined vaccine group (first dose for HB–BCG Combined vaccine, second and third dose for HB vaccine) and HB vaccine group (three dose all for HB vaccine) demonstrated that anti-HBs levels of the HB–BCG combined vaccine group were higher than that of HB vaccine group. No statistical significance was observed between the combined vaccine group and HB vaccine group after three doses immunization schedules. The results of safety in HB–BCG combined vaccine group accorded with that of BCG group, it had been not found the pathological changes of the tuberculosis. The characteristic and process in pathological changes of HB–BCG combined vaccine group and BCG group were similar in the partial poison test. HBsAg did not strengthen the inflammation reaction caused by BCG. Systemic allergy had not been found. The HB–BCG combined vaccine was stable in 2 years.

Conclusion

The immune effects of the HB–BCG combined vaccine were not lower than the two kinds of single dose vaccine, it had good safety.     

Toxicity of Tungsten Carbide and Cobalt-Doped Tungsten Carbide Nanoparticles in Mammalian Cells in Vitro

Background

Tungsten carbide nanoparticles are being explored for their use in the manufacture of hard metals. To develop nanoparticles for broad applications, potential risks to human health and the environment should be evaluated and taken into consideration.

Objective

We aimed to assess the toxicity of well-characterized tungsten carbide (WC) and cobaltdoped tungsten carbide (WC-Co) nanoparticle suspensions in an array of mammalian cells.

Methods

We examined acute toxicity of WC and of WC-Co (10% weight content Co) nanoparticles in different human cell lines (lung, skin, and colon) as well as in rat neuronal and glial cells (i.e., primary neuronal and astroglial cultures and the oligodendro cyte precursor cell line OLN-93). Furthermore, using electron microscopy, we assessed whether nanoparticles can be taken up by living cells. We chose these in vitro systems in order to evaluate for potential toxicity of the nanoparticles in different mammalian organs (i.e., lung, skin, intestine, and brain).

Results

Chemical–physical characterization confirmed that WC as well as WC-Co nanoparticles with a mean particle size of 145 nm form stable suspensions in serum-containing cell culture media. WC nanoparticles were not acutely toxic to the studied cell lines. However, cytotoxicity became apparent when particles were doped with Co. The most sensitive were astrocytes and colon epithelial cells. Cytotoxicity of WC-Co nanoparticles was higher than expected based on the ionic Co content of the particles. Analysis by electron microscopy demonstrated presence of WC nanoparticles within mammalian cells.

Conclusions

Our findings demonstrate that doping of WC nanoparticles with Co markedly increases their cytotoxic effect and that the presence of WC-Co in particulate form is essential to elicit this combinatorial effect.     

The Use of Recombinant 31 kDa Antigens of Trichinella spiralis for Serodiagnosis of Experimental Trichinellosis

Background:

We have previously reported that a 31 kDa protein was screened from the excretory-secretory (ES) proteins of Tichinella spiralis muscle larvae (ML) by immunoproteomics using early infection sera, and the gene encoding a 31 kDa protein from T. spiralis was cloned and expressed in an E. coli expression system. In this study, the recombinant 31 kDa antigens were used for detection of anti-Trichinella antibodies in serum of experimentally infected mice by ELISA.

Methods:

Anti-Trichinella IgG antibodies in sera of mice infected with Trichinella were assayed by ELISA with recombinant 31 kDa antigens, and its sensitivity and specificity were compared with ELISA with ES antigen.

Results:

The sensitivity and specificity of ELISA with recombinant antigens was 96.67% (29/30) and 96.87% (62/64), compared with 100% (30/30) and 98.44% (63/64) of ELISA with ES antigens was (P>0.05). In heavily, moderately and lightly infected mice (500, 300 and 100 larvae/mouse), anti-Trichinella antibodies were firstly detected by ELISA with recombinant antigens at 8, 12 and 14 dpi, respectively; then increased rapidly with a detection rate of 100% respectively at 28, 22 and 30 dpi. While the antibodies were firstly detected by ELISA with ES antigens at 10, 8 and 10 dpi, respectively, the antibody positive rate reached 100% at 14, 12 and 22 dpi, respectively.

Conclusion:

The recombinant 31 kDa antigens of T. spirali had a good sensitivity and specificity for detecting anti-Trichinella antibodies and might be the potential diagnostic antigen for trichinellosis.     

DDT Exposure of Zebrafish Embryos Enhances Seizure Susceptibility: Relationship to Fetal p,p′-DDE Burden and Domoic Acid Exposure of California Sea Lions

Background

California sea lions have a large body burden of organochlorine pesticides, and over the last decade they have also been subject to domoic acid poisoning. Domoic acid poisoning, previously recognized in adult animals, is now viewed as a major cause of prenatal mortality. The appearance of a chronic juvenile domoic acid disease in the sea lions, characterized by behavioral abnormalities and epilepsy, is consistent with early life poisoning and may be potentiated by organochlorine burden.

Objective

We investigated the interactive effect of DDT (dichlorodiphenyltrichloroethane) on neurodevelopment using a zebrafish (Danio rerio) model for seizure behavior to examine the susceptibility to domoic acid–induced seizures after completion of neurodevelopment.

Methods

Embryos were exposed (6–30 hr postfertilization) to either o,p′-DDT or p,p′-DDE (dichlorodiphenyldichloroethylene) during neurodevelopment via a 0.1% dimethyl sulfoxide solution. These larval (7 days postfertilization) fish were then exposed to either the seizure-inducing drug pentylenetetrazol (PTZ) or domoic acid; resulting seizure behavior was monitored and analyzed for changes using cameras and behavioral tracking software.

Results

Embryonic exposure to DDTs enhanced PTZ seizures and caused distinct and increased seizure behaviors to domoic acid, most notably a type of head-shaking behavior.

Conclusion

These studies demonstrate that embryonic exposure to DDTs leads to asymptomatic animals at completion of neurodevelopment with greater sensitivity to domoic acid–induced seizures. The body burden levels of p,p′-DDE are close to the range recently found in fetal California sea lions and suggest a potential interactive effect of p,p′-DDE embryonic poisoning and domoic acid toxicity.     

Marked liver tumorigenesis by Helicobacter hepaticus requires perinatal exposure

Background

Although severe hepatitis and liver tumors occur in a high percentage of A/J male mice naturally infected with Helicobacter hepaticus, these effects have not been observed after injection of adult mice with the bacteria.

Objectives

We tested the hypothesis that perinatal exposure to the bacteria is required for liver tumorigenesis.

Methods

A/J female mice were infected by intragastric (ig) or intraperitoneal (ip) treatment with 1.5 × 10⁸ H. hepaticus before pregnancy. We examined offspring at progressive time intervals, including some kept until natural death in old age. A/J, BALB/c, and C57BL/6 weanling male mice were similarly treated ig with the bacteria and observed for up to 2 years.

Results

After ip bacterial infection of A/J females, 41% of their male offspring developed hepatitis and 33% had hepatocellular tumors, including 18% with hepatocellular carcinoma. Treatment by the ig route resulted in a similar incidence of hepatitis in offspring (35%) but fewer total liver tumors (8%) and carcinomas (4%). By contrast, ig instillation of H. hepaticus in weanling A/J, C57BL/6, or BALB/c mice resulted in low incidence of hepatitis (0–20%) and few liver tumors, despite presence of bacteria confirmed in feces.

Conclusions

Results indicate that a high incidence of liver tumors in mice infected with H. hepaticus requires perinatal exposure. Contributing perinatal factors could include known high sensitivity of neonatal liver to tumor initiation, and/or modulation of immune response to the bacterium or its toxins. Mechanisms of human perinatal sensitivity to such phenomena can be studied with this model.     

Occupational exposure to Aspergillus and aflatoxins among food-grain workers in India

Background:

Aflatoxins are a metabolite of Aspergillus molds and are widespread in the natural environment. Workers who handle food grains are at increased risk of exposure to aflatoxins and subsequently certain respiratory conditions. In India, more than half of the employed population is engaged in some type of agricultural work, yet little known about the respiratory problems as a result of exposure to aflatoxins among workers who handle food grains in India.

Objectives:

The aim of this study was to determine the risk of occupational exposure to aflatoxins in food-grain workers compared to workers who are not occupationally exposed to food grains.

Methods:

Bronchoalveolar lavage (BAL) and serum samples from 46 food-grain workers and 44 non-food-grain workers were analyzed for the presence of aflatoxins. Microscopy and culture of BAL samples were performed to detect Aspergillus species.

Results:

Aflatoxins were detected in 32.6% of the food-grain workers and 9.1% of non food grain workers (P<0.01). A significant difference was also found in BAL culture for Aspergillus (P<0.01) between the two groups. About 47.8% of the food-grain workers and 11.4% of non-food-grain workers had chronic respiratory symptoms.

Conclusion:

Occupational exposure to aflatoxins in food-grain workers was found to be associated with the increased presence of respiratory symptoms.     

Detection of Acanthamoeba and Toxoplasma in River Water Samples by Molecular Methods in Iran

Background:

Free-living amoebae such as Acanthamoeba species may act as carriers of Cryptosporidium and Toxoplasma oocysts, thus, may play an important role in the water-borne transmission of these parasites. In the present study, a loop mediated isothermal amplification (LAMP) method for detection of Toxoplasma and a PCR assay were developed for investigation of Acanthamoeba in environmental water samples.

Methods:

A total of 34 samples were collected from the surface water in Guilan Province. Water samples were filtrated with membrane filters and followed by DNA extraction. PCR and LAMP methods used for detection of the protozoan parasites Acanthamoeba and Toxoplasma respectively.

Results:

Totally 30 and 2 of 34 samples were positive for Acanthamoeba and Toxoplasma oocysts respectively. Two samples were positive for both investigated parasites.

Conclusion:

The investigated water supplies, are contaminated by Toxoplasma and Acanthamoeba (oo)cystes. Acanthamoeba may play an important role in water-borne transmission of Toxoplasma in the study area. For the first time in Iran, protocol of LAMP method was used effectively for the detection of Toxoplasma in surface water samples in Iran.