Ziehl-Neelsen Technique Research Articles

Assessment of the validity of rapid diagnostic test kits available in the Nigerian market for <i>Mycobacterium tuberculosis</i>

Tuberculosis (TB) is an infectious disease caused by various strains of mycobacteria, usually Mycobacterium tuberculosis . Early diagnosis is important in TB disease control. The use of rapid diagnostic test (RDT) kits drastically reduces the time required for reaching clinical diagnosis and this has been successful in diagnosis of HIV, syphilis and more recently malaria to mention a few. What is the case for RDTs for diagnosis of tuberculosis? Thus, the aim of this study is to evaluate and compare the performances of results produced by different rapid diagnostic test strips available in the Nigerian market for TB. A total of one hundred and eighty-four (184) subjects aged between 12-68 years were recruited for the study. These were referred from the chest clinic with a high index of suspicion of TB. Sputum samples were collected for AFB detection using the Ziehl-Neelsen and Auramine-phenol staining techniques. Blood samples were collected for serology tests using five (5) different rapid diagnostic test kits from different manufacturers, HIV status determination and evaluation of the haematological parameters we carried out. As a result, there were significant differences in the results obtained between AFB tests and serological methods with P<0.01 in all cases. The sensitivity and specificity respectively of the five different kits were 28.6% and 32.0% (Nova), 19.5% and 29.2% (Fistech), 9.1% and 27.1% (Diaspot), 12.4% and 27.7% (Abcon) and 12.4% and 27.7% (Global).finally, findings from this study show that these rapid serological tests are poor in diagnosing tuberculosis and cannot be recommended for use in this environment. © 2015 International Formulae Group. All rights reserved. Keywords: Tuberculosis, rapid diagnostic tests, acid fast bacilli

Frequency of Rapid Growing Mycobacteria among Tuberculosis Suspected Patients in Basra- Iraq

Objective: The purpose of this study was to estimate the frequency of rapid growing mycobacteria among tuberculosis suspected patients in Basra governorate and study their resistance to drugs. Methods: A total of 150 sputum samples were obtained from 150 suspected patients who attended the Advisory Clinic for Chest Diseases and Respiratory (ACCDR) in the Basra Governorate from 01/03/2013 to 1/02/2014. Smears were stained with the Ziehl Neelsen technique and specimens were inoculated on Lowenstein Jensen medium, Identification to species level was achieved on the basis of the growth characteristics, pigment production and conventional biochemical tests. Drug susceptibility was tested to rifampicin, ethambutol, pyrazinamide, isoniazid, and streptomycin using the proportional method. Results: From 150 sputum samples, 23 isolates were Mycobacterium tuberculosis (MTB) (15.33%) and 16 (10.66%) were nontuberculous mycobacteria, seven isolates of them (43.75%), 2 males and 5 females, mean age 40 years, were identified using biochemical tests as rapid growing mycobacteria of which 4 (25%) as M. chelonae, 2 (12.5%) M. abscessus and 1 (6.2%) M. smegmatis. In addition to that, the bacteria were successfully differentiated by Duplex-PCR, as MTB and NTM based on amplification of rpoB gene sequences. Sequencing of 16S rDNA showed matching with 6 biochemically identified ones, and one of the 4 M. chelonae was M. chitae. Drug susceptibility testing showed that one M. abscessus isolate appeared to be resistant to all antibiotics (TDR), while two isolates of M. chelonae showed resistance to ethambutol and rifampicin, while M. smegmatis showed weak resistance to pyrazinamide and resistance to rifampicin. Also, all isolates of M. chelonae were sensitive to pyrazinamide, isoniazid and streptomycin. Conclusion: It appears that the rapidly growing mycobacteria represents a high frequency, among nontuberculosis detected patients, which requires phenotypical and genotypical confirmation on a follow-up, along with the examination of patterns of sensitivity. Implementing Duplex-PCR proved to be decisive for differentiating NTM from MTB.

Comparative study of clinico-bacterio-radiological profile and treatment outcome of smokers and nonsmokers suffering from pulmonary tuberculosis.

Introduction:Tuberculosis (TB) is one of the leading causes of death and disease worldwide. Tobacco smoking has been linked as a risk factor for TB. This study was aimed to affirm the strength of association between smoking and pulmonary TB.Materials and Methods:Pulmonary TB patients aged between 18 and 65 years were enrolled and followed-up until treatment completion. Two consecutive sputum smears were examined from each patient for the presence of acid-fast bacilli (AFB) using Ziehl–Neelsen technique. Radiological severity of disease was assessed using guidelines of National TB Association of USA. Sputum smears for AFB were graded for positivity as per WHO Revised National TB Control Programme criteria. Response was determined in terms of sputum conversion at the end of intensive phase and final treatment outcomes.Results:Sputum smear grading of 3+ increased from 12.5% to 68.18% and 66.66% as smoking index increased from <100 to 100–299 and >300 (P < 0.05). In nonsmokers, 79.2% patients had minimal disease while only 4.2% had advanced disease as compared to smokers where 52.4% had moderate disease, 26.2% advanced disease, and 21.4% minimal disease (P < 0.01). Smokers had significantly lower treatment success rate (69%) as against nonsmokers and former smokers (93.8% and 90.9%, respectively, P = 0.001) owing to a higher default rate among smokers (28.5%) than nonsmokers (6.3%) and former smokers (9.1%).Conclusion:Smokers during initial presentation, as well as at end of the treatment demonstrate more radiological findings, cavitary disease, and worse sputum AFB smear grading. Smokers also have a poorer treatment success rate largely due to high percentage of default rate thus suggesting noncompliance as a main confounder to treatment success. Focus needs to be made to reduce defaulters which are more common among smokers.

Diagnostic Accuracy of Xpert MTB/RIF Assay in Diagnosis of Pulmonary Tuberculosis

Objectives/Background: The objectives of this study are to assess diagnostic of Xpert MTB/RIF assay for pulmonary tuberculosis (PTB) diagnosis and to compare it with microscopy (Ziehl-Neelsen staining) in the detection of pulmonary TB in Anambra state, Nigeria. Methods: A total of 1500 individuals with presumptive PTB participated. Participant’s age ranged from 15 years and above. Two sputum specimens were submitted on-the spot (S-S) within an hour interval. Sputum specimens were processed directly from Xpert MTB/RIF test according to manufacturer’s protocol whereas sputum smears were stained using Ziehl-Neelsen technique. Results: Xpert MTB/RIF test detected Mycobacterium tuberculosis (MTB) in 389 (25.9%) out of 1500 sputum specimens while microscopy detected only 237 (15.8%). Xpert MTB/RIF thus detected all the 237 smear positive cases of microscopy in addition to 152 (12.0%) positive cases among 1263 smear negative cases. Among 389 specimens with MTB detected by Xpert, Rifampicin resistance was seen in 12 (3.1%) cases. Most of the PTB patients were young men whose occupation was trading. Conclusion: Xpert MTB/Rif assay is an accurate method for rapid diagnosis of PTB. It outperformed microscopy and detected MTB in smear negative cases. It could therefore be used as an initial diagnostic test for PTB detection and rifampicin resistance in Anambra and Nigeria in general.

Risk Factors Associated with Parasitic Infection Among Municipality Solid-Waste Workers in an Egyptian Community.

Solid-waste management is associated with several health hazards, particularly parasitic infection. The objective of the study was to determine the association between risk factors and the occurrence of intestinal parasitic infections (potentially pathogenic) among municipal waste collectors in Alexandria, Egypt. A cross-sectional survey was conducted in the main municipality company in Alexandria. A total of 346 municipality solid-waste workers (MSWWs) was interviewed using an in-depth questionnaire. The type of parasitic infections among waste handlers was determined using formol-ether concentration and modified Ziehl-Neelsen technique. About half of the workers were infected with parasites. The profile of parasitic infection revealed 12 parasitic species. These were comprised of the following helminths: Schistosoma mansoni (13.3%), Enterobius vermicularis (1.7%), Ascaris lumbricoides (1.4%), and Hymenolepis nana ova (0.6%). Among protozoa were pathogenic Entamoeba histolytica (3.2%), Giardia intestinalis (2.9%), nonpathogenic protozoa such as Entamoeba coli (1.7%), and potentially pathogenic or opportunistic ones as Cryptosporidium (23.4%), Microsporidia (20.25%), Cyclospora (2.0%), Blastocystis hominis (1.7%), and Cystoisospora belli (1.2%). About 1.4% of MSWWs have pediculosis and phthiriasis in their scalp and eyelashes respectively. Risk factors for infection were associated with direct exposure to solid fecal waste (odds ratio [OR] = 1.8, confidence interval [CI] = 1.1-3.0) and occupational activities that allowed for direct exposure to solid fecal waste (OR = 2.3, CI = 1.4-4.0). Logistic regression model has revealed that educational level and residence were the factors that contribute to parasitic infection among MSWWs (P < 0.05). MSWWs are at high risk of acquiring parasitic infections. Data of the present study highlighted the need for greater biomonitoring of MSWWs and the improvement of environmental conditions and health care in such marginalized communities to prevent parasitic infection and associated morbidities.

Prevalence of intestinal protozoan infections among individuals living with HIV/AIDS at Felegehiwot Referral Hospital, Bahir Dar, Ethiopia

HIV infection continues to pose a serious challenge to global health by predisposing patients to opportunistic infections. A cross-sectional study was conducted from December 2012 to February 2013 to assess the enteric protozoan infection status among individuals living with HIV/AIDS in Felegehiwot Referral Hospital, Bahir Dar, northwest Ethiopia. Stool specimens from 399 HIV-positive individuals were examined for the presence of trophozoites, cysts, and oocysts using direct wet mount, formol-ether sedimentation and modified Ziehl-Neelsen techniques. In addition, CD4+ T-cell counts were measured to evaluate the immune status of the study subjects. An overall prevalence of 30.6% enteric protozoan infections was recorded. Pre-ART (antiretroviral treatment) individuals were more infected than patients on ART, although this was not statistically significant (p>0.05). The highest prevalence of enteric protozoan infection was due to Entamoeba histolytica/E. dispar (19.3%), followed by Cryptosporidium spp (5.8%), Giardia lamblia (4.3%), and Isospora belli (1.3%). A CD4+ T-cell count of <200 cells/μl and status of being diarrhoeic were significantly associated with the overall prevalence of enteric protozoan infection. A relatively high prevalence of enteric protozoan infection was observed among individuals living with HIV/AIDS. Routine stool and CD4+ T-cell examinations should be conducted to monitor the status of HIV/AIDS patients.

Detection of Mycobacterium bovis in Organs of Slaughtered Cattle by DNA-Based Polymerase Chain Reaction and Ziehl-Neelsen Techniques in Bauchi State, Nigeria.

Bovine tuberculosis is a chronic, infectious, and contagious zoonotic disease of domestic animals, wild animals, and humans. It also poses a public health threat and economic losses. This study was aimed at determining the prevalence of bovine tuberculosis in slaughtered cattle, based on PM meat inspection, Ziehl-Neelsen staining, and PCR techniques in Bauchi State, Nigeria. A Prospective study was conducted on 800 cattle slaughtered in the three Zonal abattoirs of Bauchi State, Nigeria. One hundred and twenty (15%) tissues from different organs had suspected bTB lesions at PM. Out of the samples examined 35 (29.2%) were AFB positive by ZN and 10 (8.3%) were confirmed positive for M. bovis by PCR, with an overall prevalence of 29.16% and 8.33%, respectively. Female had a higher prevalence rate than male cattle at 16.66% and 12.5 % by ZN and 5.00% and 3.33% by PCR, respectively (P>0.05, χ 2 = 0.218). However, there was a statistically significant association (P<0.05, χ 2 = 7.002) between detection of bTB and the age of cattle. ZN revealed that cattle aged 6 years and above had the highest number of positive bTB cases 67.9%, while cattle aged 3–5 years had the lowest 14.81%. PCR technique revealed that the cattle aged 6 and above years also had the highest percentage positive M. bovis cases of 22.84%, whereas cattle aged 3–5 years had the lowest and the overall prevalence rate of 8.33%. The study found a high infection rate of bTB among cattle and majority of the lesions 54.2% were from lungs. The prevalence of bTB was higher in Bauchi metropolitan abattoir which supplies larger population of the state with beef.

Identification and Susceptibility Testing on Samples of Mycobacterium spp

Background: Tuberculosis is still a major global health problem. Human tuberculosis is caused by species of bacteria belonging to the Mycobacterium genus. In this study we determined mycobacterial species affecting patients from Botucatu, Brazil, and tested M. tuberculosis sensitivity to different drugs. Methods: Data were obtained from Clinical Laboratory Analysis records at Botucatu Medical School University Hospital, UNESP. All samples were processed according to standard isolation procedures from the 2008 Brazil Ministry of Health Mycobacteria Manual, which consist of staining smears by the Ziehl-Neelsen technique and seeding cultures in the Lowenstein-Jensen medium. Results: Samples were isolated from sputum (80.5%), bronchoalveolar lavage (13.8%), pleural fluid (4.6%), and cerebrospinal liquor (1.1%). Smears were evaluated in 87 cases and a total of 59 Short Research Article Calsolari et al.; BJMMR, 7(11): 942-947, 2015; Article no.BJMMR.2015.408 943 patients showed positive smears; 55 from 70 sputum samples and 4 from 12 bronchoalveolar lavage samples. No pleural fluid (4) or cerebrospinal liquor (1) samples showed positive smears. The most commonly identified strain was M. tuberculosis (61 cases); followed by M. avium and M. gordonae 2 cases each, and M. peregrinum and M. abscessus 1 case each. Mycobacteria were not identified in 20 patients. Only two strains of M. tuberculosis were multidrug resistant; one was resistant to isoniazid, rifampicin, and pyrazinamide. These two patients evolved to cure. Conclusion: This study highlights a small but troubling percentage of multidrug resistant samples and reveals the occurrence of nontuberculous mycobacteria, emphasizing the importance of correctly identifying species and testing sensitivity to antibacilar drugs to assure an adequate therapy.

Coexistence of opportunistic mycosis and mycobacterium tuberculosis in patients attending the Central Tuberculosis Reference Laboratory of Ghaemshahr city, Iran

IntroductionOpportunistic fungal organisms, such as Aspergillus and Candida species, tend to cause diseases in tuberculosis (TB) patients. Respiratory tract mycosis has clinical and radiological characteristics which are very similar to TB, thereby making the disease easily misdiagnosed. TB can be a predisposition to serious opportunistic fungal infections. Prolonged use of anti-TB drugs promotes the growth and reproduction of opportunistic fungi and, in turn, aggravates the course of the underlying process in the lung tissues. The study objective is to determine fungal coexistence with mycobacterium tuberculosis (MTB) in patients attending the Central Tuberculosis Reference Laboratory of the Health Center in Ghaemshahr city, Iran. Material and methodsTwenty-five participants were recruited into the study during two months. For each patient, three successive morning samples during one month were collected into sterile wide-neck universal sputum mugs. Specimen collection was taken for analysis at the Laboratory. None of the participants in this study had been placed on anti-fungal therapy. Sputa samples were studied for fungal elements and MTB. Direct microscopy by three stainings – KOH+Calcofluor white, Gomori-methenamine-silver and Giemsa – and fungal culture were done on Sabouraud Dextrose agar. Analysis for acid-fast bacilli (AFB) was done by the Ziehl–Neelsen technique. ResultsA total of 75 fresh sputa samples were collected. Branching hyphae in 10 samples of 5 (20%) patients and pseudo-hyphae and blastoconidia in 12 samples of 5 (20%) patients were seen together with TB. Aspergillus flavus and Candida albicans were identified in the culture method and mycological exams. DiscussionMycotic disease is an important coexistence in patients with TB. Although active mycosis may be an independent marker of advanced immunosuppression, it may also act as a co-factor in accelerating and amplifying the clinical course of TB disease. It is necessary in TB patients to increase the cure rate considering coexistencing opportunistic infections.

Isolation and Identification of &lt;i&gt;Mycobacterium chelonae&lt;/i&gt; from Human Sputum among Suspected Pulmonary Tuberculosis Infections in Basra-Iraq

Objective: The purpose of this study was to estimate the frequency of Mycobacterium Chelonae (M. chelonae) among tuberculosis suspected patients in Basra governorate and evaluate the antimicrobial susceptibility. Methods: Isolation of M. chelonae from 150 samples from patients attended to the Advisory Clinic for Chest Diseases and Respiratory (ACCDR) in Basra city, smears were stained with the Ziehl Neelsen technique. Specimens were inoculated on Lowenstein Jensen medium, Identification to species level was achieved on the basis of the growth characteristics. Drug susceptibility were tested to antibiotics (Rifampicin, Isoniazid, Ethambutol, Pyrazinamide and Streptomycin) using proportional method. Results: from 150 sputum samples among tuberculosis suspected patients, sixteen samples (10.6%) were Nontuberculosis mycobacterium (NTM), from 16 NTM samples, 4 (2.6%) were M. chelonae. Drug susceptibility results showed that all isolates resistance to rifampicin, while one isolate showed intermediate resistance to ethambutol. All isolates of M. chelonae were sensitive to pyrazinamide, isoniazid and streptomycin. Conclusion: The M. chelonae present a high frequency, especially among tuberculosis suspected patients, which requires confirmation on a follow-up, along with the examination of patterns of sensitivity, is an absolute necessity in health centers in Iraq.

Same day sputum smear microscopy for the diagnosis of pulmonary tuberculosis: Ziehl-Neelsen versus fluorescent staining.

Background:Sputum smear microscopy is the main tool for the diagnosis of pulmonary tuberculosis (TB), especially in low- and middle-income countries (LMICs). Limited sensitivity of smear microscopy and patient dropouts (PDs) are the important obstacles of national TB control programs.Objectives:(1) To assess the diagnostic utility of the same day (SS2) approach (2) To compare the smear results of the spot morning (SM) and the SS2 approaches.Materials and Methods:The study was conducted in the Department of Microbiology, GSL Medical College, Rajahmundry, Andhra Pradesh, India from January 2011 to February 2015. Three sputum samples were collected [spot (S), second spot (S2) 1 h after S, and morning sample (M)] from the volunteers. The sputum smears were stained by Ziehl-Neelsen (ZN), modified ZN (MZN), and fluorescent staining (FS) techniques and the results were pooled and compared under SM and SS2 approaches.Results:Of the 3,186 study participants, sputum smear positivity (SSP) for SM approach was 9.6% and 10.8% and for SS2 approach, it was 9.4% and 10.6%, respectively, with ZN and FS and the results were statistically insignificant (Mann-Whitney U test, P > 0.05).Conclusion:Technically SSP was similar for both the approaches and no improvement was observed with the SS2 approach. Hence, there is an urgent need to improve SSP.

An unusual presentation of leprosy: Lepromatous lymphadenitis

Leprosy is a nonfatal, chronic inflammatory disease caused by Mycobacterium leprae , whose clinical manifestations are largely confined to the skin, peripheral nervous system, upper respiratory tract, eyes, and testes. However, here we are reporting a case of leprosy in a 45-year-old male, who presented with bilateral inguinal lymphadenopathy. Left inguinal lymph node was excised surgically and was transported in 10% formalin. Sections were stained with Hematoxylin and Eosin and with modified Ziehl-Neelsen (ZN) technique and were examined under light microscope. Microscopic examination revealed aggregates of foamy histiocytes surrounded by plasma cell infiltrate. Modified ZN stain was performed which showed foamy histiocytes packed with acid-fast lepra bacilli. This report highlights the importance of systemic involvement in leprosy, especially the unusual presentation with lymphadenopathy. Hence, clinicians practicing in endemic areas should bear this fact in mind.

Mycobacterium chelonae isolates among new and previously treated pulmonary tuberculosis patients in Basra Government, South of Iraq, and anti-tuberculosis drugs resistance

ObjectiveThe purpose of this study is to estimate the frequency of Mycobacterium chelonae among tuberculosis suspected patients in Basra Governorate and to evaluate the antimicrobial susceptibility. MethodsAfter isolation of M. chelonae from 100 samples from patients admitted to the RPDC, smears were stained with the Ziehl Neelsen technique. Specimens were inoculated on Lowenstein Jensen medium. Identification to species level was achieved on the basis of the growth characteristics. Drug susceptibility was tested with antibiotics (Rifampicin, Isoniazid, Ethambutol, Pyrazinamide and Streptomycin) using proportional method. ResultsFrom the 100 sputum samples among tuberculosis suspected patients, 16 samples (16%) were Non-tuberculosis mycobacterium (NTM), 4 (4%) were M. chelonae. Drug susceptibility results showed all isolates resistant to rifampicin, while one isolate showed intermediate resistance to ethambutol. Also, all isolates of M. chelonae were sensitive to pyrazinamide, isoniazid and streptomycin. ConclusionThe results of this study emphasize that M. chelonae presents with high frequency, especially among tuberculosis suspected patients, which requires confirmation on a follow-up visit, along with the examination of patterns of sensitivity, and it is an absolute necessity in addition to the approximate hour patients typically spend in health centers in Iraq.

Age and Sex Differences in Sputum Smear Microscopy Results for Acid Fast Bacilli in a Tertiary Care Centre, South India

Background and Objectives. Low counts are more difficult to find in microscopic sputum examination and thus are more likely to be missed. In this study, we aimed to estimate the proportion of low-count grading and assessing any age and gender differences in sputum smear grading in a low HIV prevalence setting. Materials and Methods. From the tuberculosis laboratory register information on sputum positivity including the grading of smears, age and gender were extracted for January 2011–December 2011. Smears were examined using Ziehl-Neelsen technique and graded as per the Program Guidelines. Positive smears were classified into low grade positive smears (scanty and 1+) and high grade positive smears (2+ and 3+). Differences in grading of smear based on gender and age were analysed using chi square test. Results. Of 9000 smears examined, 8210 (91.2%) were collected as diagnostic smears from tuberculosis suspects. Low grade positivity was 37.5% among diagnostic smears and 69.6% among follow-up smears. Sputum smears from female examinees had higher proportions of low grade positive smears (P=0.003). Stratification of age and sex within TB suspects had clearly demonstrated the observance of higher low grade positivity among female TB suspects at extremes of age.

Recognition of Cryptosporidium oocysts in fresh and old stool samples: comparison of four techniques

To perform a comparative diagnosis of Cryptosporidium infection in fresh and old stool specimens. A total of 518 stool samples including 464 diarrheic human samples, 53 samples from calves with or without diarrhea and one sample from a sheep were screened. Initially, the auramine phenol, modified Ziehl-Neelsen and ELISA techniques, were preformed. Subsequently, PCR of the 18S rRNA gene of Cryptosporidium was carried out on all positive samples obtained by microscopy and ELISA. Auramine phenol and Ziehl-Neelsen identified Cryptosporidium oocysts in 49 (9.5%) and 56 (10.8%) out of 518 samples, respectively. Cryptosporidium specific antigen was found in 97/518 (18.7%) of the samples by the ELISA test. The 18S rRNA gene of the parasite was amplified in 79 out of 119 (66.4%) samples by PCR. Comparison of the detection rates between the diagnostic methods, i.e. PCR and ELISA, showed that there was a moderate level of agreement between these tests (Kappa=0.55). The current study suggested that PCR or ELISA methods with one of microscopy technique should be used for screening of Cryptosporidium infection in a stool sample collection.

English

Intestinal parasitic infestations (IPI) are not uncommon in immunocompetent individuals. However, human immunodeficiency virus (HIV)-infected individuals with depleted immunity have an abnormally high susceptibility to infections. This study therefore, examines children with intestinal parasites according to HIV status and degree of immunosuppression. Consecutively consenting patients aged 1 to 15 years attending the Paediatric Clinic of acquired immune deficiency syndrome (AIDS) Prevention Initiative, Nigeria, were recruited as cases, while age and sex matched HIV negative controls were recruited from Out Patient Department of Jos University Teaching Hospital. Stool samples were examined for parasites by direct wet mount, formol-ether and modified Ziehl-Neelsen technique. Levels of immunosuppression were assessed amongst HIV-positive subjects. Five hundred and ten children aged 1 to 15 years equally divided between the two cohorts were enrolled for the study. Seventy-nine had IPI, giving a prevalence rate of 15.5%; 44 (8.6%) HIV positive and 35 (6.9%) HIV-negative children. The most prevalent extracellular parasite was Giardia lamblia, however HIV positives had significantly higher rate of G. lamblia infestation. Among the intracellular parasites, the infection rate in HIV-positive subjects (5.9%) was three times that in HIV-negative subjects (2.0%). HIV positive children with advanced and severe immunosuppression had significantly higher intracellular parasites. HIV status did not significantly predict the overall risk of having extracellular intestinal parasites however, it was noted that G. lamblia infection was significantly higher in HIV positive children. HIV positive children had higher risk of having intracellular parasites especially if they have advanced or severe immunosuppression. Therefore, the policy of screening children for intestinal parasites should continue irrespective of their HIV status. Those that are HIV positive children should specifically be screened for intracellular parasites. Key words: Intestinal parasites, HIV, Children, Prevalence, Immunosuppression.

Molecular Detection of Toxoplasma gondii Oocytes in the Soil from the Public Parks of the Arak City, Iran

Background and Aim: Soil structure is mainly composed of sand, silt, c lay, and organic materials. Organisms can live in the soil. The large number of stray cats in the cities of Iran is a major environmental and hea lth problem. Toxoplasma oocysts are shed with the feces of cats, so soil is known as a potential source of transmission of toxoplasmosis. The aim of this stud y was to determine the soil contamination with Toxoplasma gondii oocyst in the public parks of the Arak city. Methods: Sixty soil samples were collected from 15 main park s of Arak city. Four soil samples from the children's playground, a potting place, around trash bins, and around toilets were taken. Oocyst was iso lated from soil by floatation in saturated sucrose. Floating debris was tested by tw o methods: staining by the modified Ziehl-Neelsen technique and PCR. The target of PCR was the 122 bp fragment of the B1 gene. Results: From 60 soil samples of public parks of Arak city, 8 samples (13%) were suspected to Toxoplasma oocyst contamination in staining smears. Only 3 samples (5%) of 60 samples were positive in PCR. The result s showed that the staining method is not a good method to detect oocysts in th e soil because the diversity of oocyst in soil is very high and similar in appearan ce. Conclusion: This study showed soils of public parks in the Arak city were contaminated to oocyst of Toxoplasma. Also molecular method for the detection of parasites in the soil was more suitable than sta ining method.

Cryptosporidium spp., comparative diagnosis and geospatial distribution in diarrheic calves from dairy farms, Valdivia, Chile

ABSTRACTObjective. To determine the Cryptosporidium spp. infection frequency by using Ziehl-Neelsen and Auramine stains on samples obtained from diarrheic calves from milking farms of the Valdivia province. To compare both diagnostic tests and to determine the geospatial distribution of the infections caused by this protozoan. Materials and methods. 221 fecal samples of diarrheic calves of 24 milking farms of the Valdivia province were studied. The processing and analysis of the samples was done by Ziehl-Neelsen (ZN) and Auramine (AU) staining techniques, and the results were compared by McNemar statistical test and the concordance level was determined by kappa index. A map was also generated to determine the geospatial distribution of Cryptosporidium infections. Results. 57.9% of all the animals tested were classified as positive with the ZN stain test, while 55.6% of all the animals turned out positive for the AU stain test. The McNemar test showed no significant difference between both diagnostic techniques (p&gt;0.05), while the kappa index showed proper concordance between tests (κ=0.73). 100% of the farms studied showed protozoan presence demonstrating the broad distribution of the parasite, however, and considering the previous factor, it was not possible to determine geospatial associations for the parasite distribution. Conclusions. The infection frequency of Cryptosporidium is higher than 50% in the milking farms studied from the Valdivia province. No difference between the Ziehl-Neelsen and Auramine staining techniques was demonstrated showing very consistent results. It was possible to detect that the number of farms infected correspond to 100% of the farms analyzed.

Cryptosporidiosis - an occupational risk and a disregarded disease in Estonia.

BackgroundCases of cryptosporidiosis have not been officially reported in Estonia after the year 2000, and the disease appears to be either under-diagnosed or under-reported.FindingsBased on a human case of cryptosporidiosis contracted during faecal sampling in dairy farms, cattle considered to be sources of infection were analysed for Cryptosporidium spp. by a modified Ziehl Neelsen technique and molecular typing. C. parvum subtype IIaA16G1R1 was detected from the human case and from calves from one of nine farms enrolled in the study providing strong circumstantial evidence of zoonotic transmission from calves to humans.ConclusionCryptosporidiosis presents an occupational risk to people with cattle contact, and may also be a risk to the human population in general. Thus increased public and medical awareness is warranted.

Performance evaluation of point-of-care test for detection of Cryptosporidium stool antigen in children and HIV infected adults.

BackgroundGastro-enteritis is associated with significant morbidity and mortality in patients with HIV/AIDS and children, and Cryptosporidium is the most important parasite implicated. To date, several commercial companies have developed simple and rapid point-of-care tests for the detection of Cryptosporidium infection; however, information is scarce regarding their diagnostic significance in Ethiopia. This study aimed at evaluating the performance of a rapid diagnostic test (RDT) for the detection of Cryptosporidium stool antigen.MethodsA hospital-based cross-sectional study was conducted in Hawassa University Hospital, southern Ethiopia from May to November 2013. Faecal samples were collected from a total of 100 children and 250 HIV infected individuals with diarrhea or CD4 T-cell count lower than 200 cells/μl. Specimens were processed using direct, formol-ether concentration and modified Ziehl-Neelsen techniques for diagnosis of Cryptosporidium and other parasites. One hundred faecal samples (50 positives for Cryptosporidium, 35 positives for other parasites and 15 negatives for any intestinal parasites) were tested using the CoproStrip™Cryptosporidium kit (Savyon Diagnostics Ltd, Israel). Test parameters were calculated using microscopy of the modified Ziehl-Neelsen stained stool smear as reference method.ResultsThe performance of the RDT was first compared to routine microscopic analysis (examination ≤10 min). The CoproStrip™Cryptosporidium RDT correctly detected 31 of 42 positive samples and 49 of 50 negative samples (i.e., 11 false negatives and 1 false positive). Sensitivity, specificity, PPV, NPV and accuracy were calculated to be 74, 98, 97, 84 and 88%, respectively. Upon thorough microscopic analysis (examination >10 min), 8 more samples with very low oocyst density were found. However, these were missed by the kit and lower the sensitivity and NPV to 62 and 72%, respectively. No cross-reactivity was observed with any of the helminthic or other protozoan parasites including Isospora and Cyclospora species.ConclusionBased on the results described herein, the CoproStrip™Cryptosporidium test could be used as an alternative to conventional microscopy especially where diagnosis of Cryptosporidium is limited due to time constraints, lack of experienced microscopists or unavailability of appropriate equipment/electricity.