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Toxoplasma gondii is an obligatory intracellular protozoan that infects all warm-blooded vertebrates. Almost one-third of people throughout the world are infected by this parasite. Although toxoplasmosis is often lethal in HIV/AIDS patients, neoplastic disease, bone marrow or heart transplant recipients, it results in life-long protective immunity in healthy people. Hence, different antigens of T. gondii such as membrane, cytoplasmic and excreted-secreted antigens (ESA) can be potential candidates for immunization. Among these antigens, ESAs play an important role in induction of immune system responses. Dense granules, micronemes and rhoptries are secretory organelles in Apicomplexa protozoa. The contents of T. gondii are factors of recognition and attachment to cells, making parasitophorous vacuole (PV), and intracellular proliferation and survival, and pathogenesis. This article reviews different kinds of ESA released from these structures. It seems necessary to identify molecular aspects of ESA before diagnosis, treatment and immunization studies.

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Background and Purpose: Toxoplasma gondii is an obligatory intracellular parasite in different cells of human beings and animals. The aim of this study was to evaluate presence and movement trend of T. gondii tachyzoites in different tissues of Balb/c, after immunization with Excretory Secretory Antigens (ESA). Materials and Methods This experimental survey has been performed on 24 Balb/c mice in case and control groups. For immunization of mice, two times, intervals two weeks, case group (n=12) received 40 µL ESA+40µL Adjuvant and control group got 40 µL PBS+40µL Adjuvant. Two weeks after the second immunization, mice were challenged with 1×104 alive and active tachyzoites of T. gondii RH strain and on 1, 2, 3 days and the last day (before death), after challenge different tissues of 3 mice from each group were prepared and stained with Giemsa stain and the slides evaluated for presence or absence of parasites and parasitic load. Results: Toxoplasma after intraperitoneal injection, in both case and control groups were able to movement to various tissues. In the case group receiving Excretory Secretory Antigens (ESA), parasitic load in spleen, liver and heart was less than control group. Conclusion: Hence, ESA reduced the parasitic load, but could not inhibit the distribution and presence of Toxoplasma in different tissues.

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Background and purpose: Toxoplasma gondii is a protozoon with worldwide distribution. In spite of increasing information about its biology, treatment of toxoplasmosis is restricted to a few drugs and unfortunately using of each of drugs is associated with significant side effects in patients. It seems that investigation on alternative drugs is necessary. Materials and methods: In case group, concentrations of 1, 5 and 10 µg/ml of ketotifen were prepared and added to RPMI (Roswell Park Memorial Institute) medium containing peritoneal macrophages. After 60 minutes, incubation and adding Toxoplasma gondii tachyzoites to medium, the inhibitory rate of different concentrations of ketotifen on entrance of Toxoplasma tachyzoites into macrophages in 30 and 60 minutes were evaluated. Control group did not receive ketotifen. Results: The best efficacy of ketotifen in inhibition of entrance of Toxoplasma tachyzoites into macrophages was observed in 10 µg/ml and in 60 minutes (72.90 ± 1.70) (P < 0.05). Conclusion: The findings of this research show that ketotifen is a suitable drug for inhibiting the entrance of Toxoplasma gondii tachyzoites into nucleated cells in vitro.

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Background and purpose: Toxoplasmosis is a common parasitic disease throughout the world and ‎one-third of the population has antibodies to Toxoplasma gondii. This disease causes serious medical ‎problems in fetuses and immunocompromised individuals. As gene encoding protein GRA14 can be ‎considered as a suitable target for DNA vaccine and designing diagnostic kits the aim of this study was to do ‎cloning and sequencing the gene encoding GRA14 protein of Toxoplasma gondii RH strain.‎ Materials and methods: DNA extraction was performed on harvested tachyzoites from mouse ‎peritoneal fluid, then PCR carried out and amplification products were analyzed by gel electrophoresis. ‎GRA14 gene was cloned in pTG19-T as a cloning vector, then recombinant plasmid confirmed by the colony-‎PCR and restriction enzyme digestion using HindIII and EcoRI, followed by sequencing.‎ Results: Evaluation of PCR products by agarose gel electrophoresis and analysis of nucleotide ‎sequencing of 1227 bp gene encoding the protein GRA14, revealed the complete homology with the recorded ‎sequences in the gene bank. Furthermore, cloning of GRA14 gene in pTG19-T vector was confirmed with ‎colony PCR and restriction enzyme digestion.‎ Conclusion: The results showed that the GRA14 gene was successfully cloned into the pTG19-T ‎vector and this plasmid can be used to design DNA vaccines and diagnostic kits in further studies.‎

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Background and purpose: Trichomoniasis, is a protozon infection in lower urinary reproductive tract with various prevalence rate in different populations. In this study we investigated that as a neglected parasitic disease in Women's Clinic attending Teaching Hospitals affiliated with Mazandaran University of Medical Sciences. Material and methods: This descriptive study was performed in 1900 samples during 2006-2014. Active and passive case finding was conducted at 400 patients and 1500 archived smears, respectively. After preparing a demographical check list, the wet smears of vaginal specimens and Pap smears were microscopically examined. For molecular confirmation of positive samples the nested PCR was performed. To analyze the data chi-square test was applied in using SPSS V.19. Results: A total of 1900 samples were analyzed in this study of which 21 (1.1%) were infected with Trichomonas vaginalis. The 1100 bp band corresponding actin gene T. vaginalis was successfully amplified. Also, 64.7% of positive samples had trichomoniasis، candidiasis and bacterial co-infections. The prevalence of infection was higher in those aged 35-44 years of old. Conclusion: The results showed that prevalence of Trichomonas vaginalis was relatively low in studied area. But, the disease is regarded as neglected among women aged 35 to 44 years of old. Therefore, further investigations are recommended among this age group.

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Background and purpose: Polycyclic aromatic hydrocarbons (PAHS) are environmental pollutants that are caused by human activity and considered as carcinogenic and mutagenic compounds. Anthracene is a hazardous substance causing serious health problems. There are many bacteria with the ability to remove this pollutant. The aim of of this study was isolation of some bacteria from Tajan river estuary and their use in biodegradation of Anthracene. Materials and methods: In this study the samples taken from estuaries were inoculated into a synthetic medium culture. Four species of bacteria were isolated in the culture process and applied for removal of Anthracene in different environmental conditions such as pH, time, temperature and concentration of Anthracene. Spectrophotometer and HPLC were used to study the growth of bacteria and Anthracene concentration, respectively. Results: The results showed that, optimum condition for removal of Anthracene was pH= 7, temperature= 30 °C, and the dose of inoculated bacteria 107 cfu/ml. Anthracene concentration was 100 ppm. Efficiency of the Anthracene removal in the presence of Pseudomonas putida, Achrobactrum haematophilum, Klebsiella oxytoca, Enterococcus sp, was obtained 51, 45, 43 and 48 percent, respectively. Pseudomonas putida bacteria species had the highest efficiency in Anthracene removal and Enterococcus sp strains had the lowest efficiency. Conclusion: The biological method is a cheap and effective method that can be used for Anthracene removal.

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Background and purpose: Fasciola hepatica and Fasciola gigantica are common liver flukes that are the etiological agents of fasciolosis, which affects both domestic livestock and humans worldwide. In the present study we established a rapid, easy and also accurate tool, for differentiation between F. hepatica and F. gigantica using Fast PCR. Material and methods: Thirty adults of Fasciola species were isolated from sheep and cattle liver form abattoirs in Mazandaran province. ITS1 rDNA region were amplified by Sapphire Amp® Fast PCR and compared with AccuPower® Taq PCR PreMix from Bioneer. In addition, PCR-RFLP assay using Tsp509I was performed for identification of F. hepatica and F. gigantica. Results: A fragment of approximately 463bp was amplified in all of the Fasciola samples using Sapphire Amp® Fast PCR premix in just 34 minutes, while nucleic acid amplification was completed in about 1 hour and 46 minutes by Bioneer PCR master mix. All PCR products were digested with restriction enzyme TasІ (Tsp509I). After digestion, F. hepatica revealed two fragments of 151 and 312 bp while F. gigantica produced three fragments of 93, 151, and 219 bp. Conclusion: Fast PCR reaction using Sapphire Amp® Fast PCR premix was completed three times faster than the time of conventional premix. The new Fast PCR assay using Sapphire Amp® Fast PCR premix provides a simple, rapid and accurate technique for identification and differentiation of Fasciola species in epidemiological researches on human and domestic animals in endemic regions of fasciolosis.

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Background and purpose: Dicroceliasis is a common disease in ruminants caused by various species of Dicrocoelium. This parasite is seen communally in bile ducts and gallbladder of ruminants and sometimes accidentally in humans. The parasite is considered to be important from both economic and veterinary aspects since it has a direct impact on liver damage and results in significant loss of protein in human diet because infected livers are removed in slaughterhouses. This parasite is prevalent in Iran, so, this study was performed to determine the rate of dicrocoeliasis infection in Iran.

Materials and methods: We sought to conduct a systematic review of articles published on some databases such as Google Scholar, PubMed, Science Direct, IranMedex, Scopus, SID, IranDoc and Magiran, between 2000-2015.

Results: Our search resulted in a total of 15 reports published about the prevalence of dicrocoeliasis infection. The random effect model was used for this meta-analysis. The relative prevalence rate of dicrocoelium was 3.1% (2.2-4.2%) in sheeps, 1.3% (0.9-1.9%) in goats, and 2.1% (1.1-3.5%) in cows.

Conclusion: This study revealed high prevalence of dicrocoelium dentriticum infection among domestic animal in Iran, therefore, it is necessary to follow hygiene procedures in washing vegetables and adequate monitoring are also needed in Iranian abattoirs to ensure the infected livers are removed.

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Background and purpose: Fasciola hepatica and Fasciola gigantica are common liver flukes which affect both human and livestock worldwide. In this study we evaluated the loop-mediated isothermal amplification (LAMP) assay for detection and discrimination of Fasciola species.

Materials and methods: Fifty adults of Fasciola worms were isolated from sheep and cattle liver form abattoirs in Mazandaran province. A total of 8 primer sets for LAMP was designed to amplify the 28S ribosomal RNA gene of Fasciola sp. Conventional LAMP was carried out in a 20µI reaction mixture under isothermal condition at 60°C for 90 minutes. Amplification result was observed by monitoring the turbidity by naked-eye, using fluorescent dye and gel electrophoresis. The specificity of LAMP method for detecting Fasciola sp. was tested by amplification of Dicrocoelium dendriticum, Trichostrongylus colubriformis, and Echinococcus granulosus DNA templates. To evaluate the detection limit of LAMP assay in detecting Fasciola genus, serial dilution of the extracted DNA was used.

Results: A positive LAMP reaction by the specific primers of two species produced many bands of different sizes in 60^0C after 90 min. The optimal assay conditions were established with no reaction with other parasites’ DNA. The detection limit of this LAMP assay was 1 pg DNA/tube. The result of turbidity and fluorescent dye detection were consistent with agarose gel electrophoresis.

Conclusion: Our results demonstrated that LAMP is a rapid, cost-effective, highly specific, easy, and reliable method for differentiation of Fasciola sp. in epidemiological and clinical researches on human and domestic animals in endemic regions of fasciolosis.

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Background and purpose: Wild boars (Sus scrofa) are potential reservoirs for a lot of zoonotic diseases, so it is possible to transmit these infections to wild and domestic animals and also humans. This research aimed to survey the protozoan prevalence of Sus scrofa in Mazandaran Province, North of Iran.
Materials and methods: In a descriptive cross-sectional study a total of 21 wild boars was captured beween December 2012 and March 2014. Faeces collected from the intestinal tracts of each animal was placed in appropriate fixative. Temporary staining with Lugol’s solution was performed in order to find the protozoan cysts and trophozoites. The stool samples were further studied using sedimentation and flotation methods. Then, fixed stool smears were examined by trichrome and Ziehl–Neelsen staining.
Results: There were 21 samples of which 12 (57.14%) were found to be infected with one or more protozoan species. The prevalence of contamination with intestinal protozoan was as follows: 33.33% trophozoites and cysts of Balantidium coli, 28.57% oocysts of Eimeria spp., and 9.52% cysts of Giardia spp.
Conclusion: According to current findings, wild boars in Mazandaran province are infected with a lot of zoonotic protozoan, which create potential risks for other animals and people in the region.

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Background and purpose: In developing countries, a high prevalence of musculoskeletal disorders is reported among drivers. However, there are few researches in Iran on this problem among suburban drivers. The present study aimed at investigating the prevalence of work-related musculoskeletal disorders and its related factors in suburban drivers in Mazandaran province.
Materials and methods: This cross-sectional descriptive analysis was carried out in 1850 suburban drivers. Nordic Musculoskeletal Questionnaire and the demographic questionnaire were completed for all drivers. Logistic regression analysis was done in SPSS V22 applying Chi-square test.
Results: The highest prevalence of musculoskeletal disorders in the past 12 months was found in neck (16.3%), knees (13.9%), and back and elbow (10.7%). Other factors such as age, work experience, smoking, previous accidents, and marital status showed a significant relationship with pain in various parts of the body (P<0.05).
Conclusion: This study showed a high prevalence of musculoskeletal disorders (especially in neck and knee) among suburban drivers in Mazandaran province. Such disorders could be easily prevented by designing ergonomic seats based on Iranian anthropometric sizes, doing stretch exercises, having sufficient rest, and trainings on correct sitting.
 
 

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Background and purpose: Dicrocoeliasis is a prevalent parasite in ruminants caused by different species of Dicrocoelium spp. This parasite lives in bile duct and gallbladder of ruminants and accidentally in human. The economic significance of dicroceliasis is due to direct damage to the liver and, consequently, to the loss of significant amounts of protein. The present study aimed to investigate the morphometric and molecular (PCR and PCR-RFLP) parameters of Dicrocoelium species isolates from sheep, goat, and cattle in Mazandaran province, Iran during 2013-2015.
Materials and methods: In this descriptive study, 50 trematodes isolated from livers of sheep (n= 20), goat (n= 20), and cattle (n= 10) were collected form abattoirs in Mazandaran province. Adult worms were studied by morphometric and molecular methods on 28s rDNA. The RFLP technique was used to identify the genus. Some samples were also sequenced.
Results: All isolates of three hosts (sheep, goat, and cattle) showed significant differences in many morphological parameters. According to characterization of testicles that is a remarkable characteristic for differential diagnosis of Dicrocoelium species, they were in tandem position in all isolates. Moreover, no significant difference was observed in their size in different hosts. Four bands, including 116, 145, 393, and 409 bp were produced from PCR method after treatment by enzyme. In molecular technique, there were no any significant differences but in morphometric analysis, some differences were found.
Conclusion: The molecular and morphometric results indicated that D. dendriticum is the only species infecting sheep, goat, and cattle in Mazandaran province.
 
 

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Background and purpose: Taxonomic identification of larval and adult stages of Taenia hydatigena from other Taenia species based on morphometric characteristics of the rostellar hooks in intermediate and definitive host is important in medical and veterinary parasitology. Therefore, this study aimed at identifying the larval stage of Taenia hydatigena (Cysticercus tenuicollis) based on morphological characteristics of rostellar hooks isolated from sheep and goat in Mazandaran province, Iran for the first time.
Materials and methods: This study was conducted in 100 samples of rostellar hooks of the larval stage of Taenia hydatigena isolated from sheep and goats in 2014-2015. Morphological characteristics of large and small hooks in the number and size characters were determined and measured by calibrating microscopes. Data analysis was done in SPSS applying Chi-square and One-way ANOVA.
Results: The mean number of large and small hooks in goat and sheep isolates were 30.8±3. Morphometric characteristics of hooks in the shape and size were similar in both animal isolates. But significant differences were seen between native and nonnative isolates in mean width of large hooks length (P< 0.05).
Conclusion: This study identified the larval stage of Teaina hydatigena from other species of Teaina according to characteristics of rostellar hooks, which is of great significance in taxonomic studies in parasitology.
 
 

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عفونت‌های انگلی روده‌ای ازجمله بیماری‌های شایع در کودکان به‌ویژه در مدارس می‌باشد. بررسی و تعیین آلودگی دانش‌آموزان به این عفونت‌ها در مقاطع مختلف به‌ویژه در مناطق روستایی و شهری از اولویت‌های تحقیقاتی به‌منظور تشخیص و درمان در انگل‌شناسی پزشکی و سیستم بهداشتی و درمانی می‌باشد(1،‌2). از طرفی انگل‌های روده‌ای باعث اختلال در رشد ذهنی و تکامل شناختی کودکان می‌شود. مهم‌ترین و ارزان‌ترین راه پیشگیری از این بیماری‌ها شناسایی مبتلایان، درمان و آموزش بهداشت است(3،‌4). با توجه به‌این‌که مطالعه‌ای در زمینه شیوع بیماری‌های انگلی روده‌ای در مدارس ابتدایی شرق استان مازندران انجام نشده است لذا مطالعه حاضر باهدف بررسی عفونت‌های انگل‌های روده‌ای در دانش‌آموزان مدارس ابتدایی خلیل شهر، در شرق استان مازندران در سال 1397 انجام شد. در این مطالعه در مجموع نمونه‌های مدفوع ۵۰۰ دانش‌آموز از شش مدرسه ابتدایی خلیل شهر در فاصله سنی ۱۲-۷ سال مورد بررسی قرار گرفتند،
که تعداد ۲۶۴ نفر (۸/۵۲ درصد) آن‌ها پسر و تعداد
۲۳۶ نفر (۲/۴۷ درصد) دختر بودند. فراوانی آلودگی انگل‌های روده‌ای در دانش‌آموزان برحسب سن مادران دانــش‌آمــوزان نشــان داد کـه میـانگیـن سنـی مــادران دانش‌آموزان آلوده ۵۰۳/۶ ± ۰۹/۳۶ (حداکثر سن ۴۲ سال و حداقل سن ۳۰ سال) بود. شیوع آلودگی انگلـی روده‌ای در دانـش‌آموزان در مـادران خانـه‌دار و خانوارهای ۳-۲ فرزند، بیش‌تر از سایر گروه ها بوده است. از۵۰۰ دانش‌آموز موردمطالعه ۱۱نفر(۲/۲ درصد) آلوده به انگل‌های روده‌ای بودند. شیوع تک‌یاخته روده‌ای بلاستوسیستیس هومینیس ۸/۱ درصد و ژیاردیا لامبلیا ۲/۰درصد بودند. در این مطالعه سایر آلودگی‌های انگلی مانند آنتامبا هیستولیتیکا، اکسیور، هیمنولیپیس نانا مشاهده نشده است. در این مطالعه ارتباط معنی‌دار بین مصرف سبزیجات خام با آلودگی انگلی روده‌ای مشاهده شد. مقایسه شیوع آلودگی‌های انگلی روده‌ای در مدارس ابتدایی در نقاط مختلف کشور با نتایج مطالعه حاضر نشان می دهد که میزان شیوع آلودگی در مدارس ابتدایی خلیل شهر نسبت به سایر مطالعات پایین‌تر بوده است و با توجه به گذشت زمان و بالا رفتن سطح بهداشت خانواده‌ها و سطح معلومات آنان از بیماری‌های انگلی، طبیعی است که میزان ابتلا به انگل‌های روده‌ای کاهش پیدا کرده باشد. به ‌منظور کنترل و کاهش آلودگی به انگل‌های روده‌ای در جمعیت موردمطالعه، مواردی همچون آموزش بهداشت در زمینه بیماری های انگلی، شناسایی و درمان مبتلایان، همکاری بین مراکز بهداشتی درمانی و آموزش و پرورش، آزمایشات غربالگری قبل از ثبت نام در شهرستان پیشنهاد می شود.

 

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Background and purpose: There are few treatment options available for treatment of toxoplasmosis and effective drugs have serious toxic effects. In this study, the in vivo and in vitro anti-toxoplasma activities of Heracleum persicum and Foeniculum vulgare fruits essential oils were investigated.
Materials and methods: In vitro, Vero cells were incubated with different concentrations of essential oils or pyrimethamine (positive control) and the cellular viability was determined. Next, Vero cells were infected with T. gondii (RH strain) and treated with agents. Then, the CC₅₀, IC₅₀, and selectivity index (SI) were calculated. Moreover, in vivo, the effect of oils on survival times of Balb/c mice infected with T. gondii were determined.
Results: In vitro results showed that the oils exhibited less cell toxicity than pyrimethamine.
The selective index was 2.94, 6.96, and 3.06 for Heracleum persicum, Foeniculum vulgare, and pyrimethamine, respectively. Also, the infected mice treated with F. vulgare-pyrimethamine showed a better survival rate than others (P<0.05).
Conclusion: The H. persicum and F. vulgare essential oils showed anti-toxoplasmic activity in vitro and in vivo, but, combination therapy with F. vulgare and pyrimethamine showed a better survival time in mice infected with T. gondii. Therefore, F. vulgare may be a useful candidate in treatment of Toxoplasmosis. However, further studies are needed to investigate the fractions of this plant against T. gondii.

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Background and purpose: Camel breeding is common in East of Iran due to its semi-arid climate and desert conditions. It transmits some diseases to other ruminants and humans due to carrying common blood parasites between humans and animals. This work aimed at studying the presence of Trypanosoma and Piroplasmida (Babesia and Theileria) species in camels in South Khorasan province using parasitological and molecular methods.
Materials and methods: In this study, blood samples were taken from 200 camels in South Khorasan that were randomly selected in 2020. Then, thick and thin smear were prepared from the samples and stained with Giemsa staining technique and examined under a light microscope. For molecular detection of Trypanosoma, Theileria and Babesia, specific primers of each genus were used.
Results: Microscopic examination showed that out of 200 blood samples, 8 samples (4%) were infected with Trypanosoma and no samples were infected with Theileria and Babesia. According to molecular experiments, six samples (3%) had DNA of Trypanosoma evansi and five samples (2.5%) were found with Trypanosoma brucei DNA. But the DNA of Theileria and Babesia were not observed. Phylogenetic analysis showed that the sample isolated in the present study had a high genetic affinity with genotype A of Trypanosoma evansi.
Conclusion: To the best of our knowledge, this was the first study to investigate the prevalence of Surra in South Khorasan province. The presence of Trypanosoma in the blood of camels in this region signifies the need for a careful monitoring system and evaluation studies as a prerequisite for controlling blood parasitic diseases in camels in South Khorasan.
 

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Background and purpose: Giardia and Blastocystis are common parasites of humans with a wide range of hosts. The aim of this study was to identify the genotypes of Giardia intestinalis and Blastocystis hominis in individuals attending Bavi health centers in Khuzestan province, Iran.
Materials and methods: In this study, 30 positive stool samples of Giardia and Blastocystis were collected from individuals attending Bavi health centers in 2019-2020. Then DNA extraction was performed on fecal samples. PCR was performed using GDH gene for Giardia and 18S rRNA region for Blastocystis and PCR products were drawn to determine sequence type and phylogenetic tree.
Results: Sequence analysis was performed on PCR products from Giardia based on amplification of GDH gene. Based on the results of DNA sequence, all identified samples belonged to assemblage BIV. Sequence analysis was performed on PCR products from Blastocystis according to amplification of 18S rRNA gene and all identified samples belonged to ST3 subtype.
Conclusion: The study showed that the predominant genotype of Giardia in Bavi was BIV which indicates a zoonotic cycle in this region. In addition, the predominant subtype of blastocystis was previously reported in this region (ST3) often seen in human-to-human transmission cycle, although there have been reports of animal-to-human cycle. Therefore, alongside health measures to control these two parasites in this area, molecular epidemiology studies in local animals is necessary to determine the zoonotic transmission of Giardia and Blastocystis parasites in this region.

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Background and purpose: Patients admitted at the intensive care unit (ICU) need blood draws for chemistry analytes and arterial blood gas (ABG) analysis. Repeated blood drawing is one of the most common causes of anemia in these group of patients. Measurement of analytes only by ABG analyzer can be beneficial for preventing anemia. The aim of this study was to compare the biochemical analytes in venous and ABG samples in ICU patients.
Materials and methods: This cross-sectional study was performed in 223 patients hospitalized in the ICU. These patients undergoing ABG analysis and simultaneous venous sampling. The value of Ca, Na, K, Hct, and glucose were measured in venous samples with a laboratory auto-analyzer and in arterial samples with an ABG analyzer.
Results: The mean age of patients was 61.14±16.96 years. The mean difference between ABG and serum samples was 1.12 mEq/L for sodium, -0.29 mEq/L for potassium, -0.98 mg/dl for calcium, 0.33 mg/dl for glucose, and 4.39%% for hematocrit. In addition, 95% limit of agreement was -12.3 to 9 for sodium, -0.7 to 1.29 for potassium, -1.31 to 3.8 for calcium, -39.13 to 40.32 for glucose, and -13.5 to 8.5 for hematocrit.
Conclusion: Because of acceptable agreement for sodium, potassium, and hematocrit, ABG analyzer can be used in ICU patients and emergency situations. There is a remarkable bias between the ABG-measured and laboratory-measured calcium and glucose, so the calcium and glucose measurement by the ABG analyzer are not reliable.