Microscopic Examination Of Smears Research Articles

Evidence of Carriage of Antimicrobial Resistant Salmonella species of Public Health and Veterinary Significance in the Intestines of House Crows (Corvus splendens) in Tanzania

The Indian house crow, Corvus splendens (Vieillot) was introduced in Zanzibar, Tanzania by the British and immigrants from India in 1897 to help clean the town. The crow is responsible for polluting the environment, water sources and human surroundings by their droppings and the rubbish they carry. This behavior has led to concern that, the crows may be responsible for the spread of certain pathogens including Salmonella and their persistence in the environment.Given the zoonotic potential of Salmonella, the main aim of this study was to investigate the occurrence of antimicrobial resistant Salmonella infections in Indian house crows and to determine if the isolates were similar to those associated with disease in livestock or humans. Methods: Indian house crows were lured with meat and blood baits to land into the crow live-trap set at the Mabibo compound of the National Institute for Medical Research (NIMR) in Dar es Salaam city in Tanzania. A total of 100 house crows were captured, humanely sacrificed, and their small and large intestines were obtained by using aseptic techniques for microbiological investigations. Culture technique was employed to detect the presence of Salmonella in intestinal contents; and preliminary identification of the isolates was based on colonial characteristics on selective media and microscopic examination of smears following Gram staining. Confirmation of Salmonella species was done by biochemical tests. Antimicrobial susceptibility testing was done by using the disc diffusion method on Mueller Hinton agar. Eight isolates were identified by standard microbiological techniques as Salmonella spp. (6 suggestive of Salmonella gallinarum and 2 suggestive of S. Typhi). All isolates were found to be susceptible to ciprofloxacin but resistant to amoxicillin. Lower levels of susceptibility were noted for chloramphenicol and ceftriaxone. Our results demonstrate the presence of antimicrobial resistant Salmonella spp. in the Indian house crows’ population and provide an indication of potential public and poultry health risks associated with these birds in the coastal area. The occurrence of antibiotic resistant S. Typhi and S. gallinarum among Indian house crows has both veterinary and public health consequences as they may be transmitted to poultry and humans. This therefore provides further rationale for the public action on eradicating the house crows.

Microsporidia Infection of the Cornea—A Unique and Challenging Disease

Microsporidia are a diverse group of obligate, intracellular, eukaryotic, spore-forming parasites. Traditionally, these were considered as protozoans but recently have been reclassified as fungi. Microsporidia behave as opportunistic pathogens causing systemic infections. In the eye, Microsporidia cause keratoconjunctivitis, corneal stromal keratitis, scleritis, and endophthalmitis. This review shares our experiences with anterior segment infections caused by this pathogen. Keratoconjunctivitis is a common form of ocular infection caused by the parasite. Although early reports described it as occurring only in immunosuppressed individuals, it can also occur in immunocompetent individuals. The disease shows a seasonal pattern with a peak incidence during the rainy season. Although several drugs have been considered, our experience suggests that keratoconjunctivitis is a self-limiting disease. In contrast to keratoconjunctivitis, stromal keratitis is an ill-defined disease. We collected 30 cases and analyzed the various aspects of this disease. Stromal keratitis is characterized by a slowly progressive course. The corneal picture resembles herpes simplex virus stromal keratitis or fungal keratitis cases, and is characterized by deep stromal infiltrates with overlying and surrounding stromal edema and keratic precipitates. The diagnosis of Microsporidia infection is confirmed by a microscopic examination of smears from patients with ulcerative keratitis or by a histopathological examination of corneal tissues. Definitive genus identification requires the examination of specimens by electron microscopy or by molecular methods. In the absence of a definitive medical treatment, nearly all patients require surgical treatment. The confusion regarding Microsporidia is not only limited to their classification but also extends to various aspects of the corneal disease caused by them.

Molecular characterization of haemosporidian parasites (Haemosporida) in yellow wagtail (Motacillaflava), with description of in vitro ookinetes of Haemoproteus motacillae.

In studies of haemosporidian systematics and taxonomy, a combination of microscopic examination and molecular identification has been recommended. The yellow wagtail (Motacillaflava) is a frequently used species in studies of host-parasite interactions; blood parasites of this bird have been well studied using microscopic examination of blood films, but data on molecular characterization of the parasites are scattered. Here we present the first study linking several haemosporidian cyt b sequences with their morphospecies parasitizing the yellow wagtail, combining light microscopy and nested PCR methods. The lineage hYWT1 of Haemoproteus motacillae was the most prevalent parasite; it was recorded in 38% of the birds. The latter lineage and the lineages hYWT2, hYWT3, hYWT5 and hYWT6 form a well-supported clade on the phylogenetic tree and likely represent intraspecific genetic variation of H. motacillae, with genetic divergence of 0.3 - 1.5 % among these lineages. Microscopic examination of smears prepared during an in vitro experiment, which was designed for observation of developing ookinetes, showed that H. motacillae ookinetes were present 5 hours after exposure of blood to air at 18-20° C. Ookinetes of this parasite belong to a group of large Haemoproteus spp. ookinetes (19.9 µm in length on average). Illustrations of the parasites are given. Two of reported Plasmodium lineages, pCOLLI and pYWT4 are phylogenetically closely related to the widespread host generalist parasites Plasmodium relictun (cyt b lineages pSGS 1, pGRW4 and pGRW11), but it was not possible to establish this morphologically. This study contributes to the molecular identification of avian haemosporidian parasites and provides information on morphology of H. motacillae ookinetes, which is additional information assisting the microscopic identification of this species.

Molecular identification of Theileria and Babesia in sheep and goats in the Black Sea Region in Turkey

This study was carried out to investigate presence and distribution of Theileria and Babesia species via microscopic examination and reverse line blotting (RLB) techniques in sheep and goats in the Black Sea region of Turkey. For this purpose, 1,128 blood samples (869 sheep and 259 goats) were collected by active surveillance from sheep and goats in different provinces of various cities in the region in the years 2010 and 2011. Smears were prepared from the blood samples, stained with Giemsa, and examined under the light microscope for Theileria and Babesia piroplasms. The genomic DNAs were extracted from blood samples. The length of 360-430-bp fragment in the variable V4 region of 18S SSU rRNA gene of Theileria and Babesia species was amplified using the gDNAs. The polymerase chain reaction products were hybridized to the membrane-connected species-specific probes. A total of 38 animals (3.37%) including 34 sheep (3.91%) and 4 goats (1.54%) were found to be positive for Theileria spp. piroplasms in microscopic examination of smears while Babesia spp. piroplasm could not detected. Infection rates were 34.64% in sheep, 10.04% in goats, and totally 28.99% for Theileria ovis while 0.58% in sheep and totally 0.44% for Babesia ovis. However, Theileria sp. OT3 was detected in 2.65% of sheep and 2.04% of all animals; besides Theileria sp., MK had 0.58% prevalence in sheep and 0.77% in goats, with a total 0.62% with RLB. Although T. ovis and Theileria sp. MK were determined in both sheep and goats, B. ovis and Theileria sp. OT3 were observed only in the sheep. These results provide the first detailed molecular data for sheep and goat theileriosis and babesiosis in the region.

Evaluation of Ziehl–Neelsen smear for diagnosis of pulmonary tuberculosis in childhood in a rural hospital in Ethiopia

The goal of this study is to describe the experience with smear microscopy examination for acid-fast bacilli (AFB) of spontaneous sputum from children in a district hospital located in a rural zone of Ethiopia. All sputum reports of children were retrospectively reviewed from July 2007 until June 2012. During the period of study, 875 children less than 15years old were screened and 48 (5.5%, 95% confidence interval [CI]: 4.1–7.3%) were diagnosed with pulmonary tuberculosis sputum smear positive. The mean age of the children with sputum positive for AFB was significantly higher than children with sputum negative for AFB (11.4 versus 10.4) (p=0.001). Only 1 out of 47 (1.3%) children of 6years or less had sputum positive for AFB. In 13- and 14-year-old children, 8.7% had sputum positive for AFB (20 of 229) (p=0.03). Spontaneous sputum has a low diagnostic yield in childhood in low-income countries. Alternative methods should be implemented in rural areas to improve diagnosis of pulmonary TB, particularly in children less than 12years old.

A cluster of avian malaria cases in a kiwi management programme

AIM: To describe a temporal cluster of avian malaria (Plasmodium spp.) at an Operation Nest Egg™ (ONE) site in Rotorua which caused mortality in a juvenile kiwi and had high population prevalence in brown kiwi (Apteryx mantelli). METHODS: A 70-day-old wild-born captive brown kiwi was submitted for post-mortem examination to Massey University Wildlife Health Centre. Post-mortem examination and histopathology were used to determine the cause of death. Plasmodium specific PCR analysis was subsequently conducted on tissue samples and 108 individual blood samples from living kiwi from five ONE breeding sites and two rowi kiwi crèches. Positive PCR products were sequenced to identify the Plasmodium spp. isolated. Where possible, blood smear microscopic examination was used to determine the level of parasitaemia in the infected kiwi. RESULTS: Plasmodium spp. was detected in the kiwi which died and it showed histopathological evidence of disseminated protozoal infection. A high prevalence of Plasmodium was found in blood samples from kiwi concurrently residing at ONE Rotorua by blood smear microscopy (22/32, 68%) and PCR (25/32, 78%). All kiwi with positive blood smears had only a low level of peripheral parasitaemia at the time of sampling. However, 0/17 additional kiwi sampled at Rotorua 3 weeks after the juvenile's death, 0/23 Rotorua juveniles sampled 1 year later and 0/59 kiwi from the five other locations were positive for Plasmodium by these methods. Sequencing analysis revealed a cosmopolitan Plasmodium (Huffia) elongatum lineage in all positive birds. CONCLUSIONS: This is the first description of an avian malaria (Plasmodium spp.) infection associated with mortality and a high population prevalence in brown kiwi at a ONE site in the 20 years of the programme. The study suggests that this level of infection in a population of kiwi was unusual and provides evidence in support of continued vigilance of disease risks associated with this and other conservation management programmes involving wildlife translocation.

Should Sputum Smear Examination Be Carried Out at the End of the Intensive Phase and End of Treatment in Sputum Smear Negative Pulmonary TB Patients?

BackgroundThe Indian guidelines on following up sputum smear-negative Pulmonary tuberculosis (PTB) patients differ from the current World Health Organization (WHO) guidelines in that the former recommends two follow up sputum examinations (once at the end of intensive phase and the other at the end of treatment) while the latter recommends only one follow up sputum smear microscopy examination, which is done at the end of the intensive phase. This study was conducted to examine if there was any added value in performing an additional sputum smear examination at the end of treatment within the context of a national TB program.MethodsThis study was a descriptive record based review conducted in nine tuberculosis (TB) units in Delhi, India. All consecutive new sputum smear-negative PTB patients registered in these nine TB units from 1st January 2009 to 31st December 2009 were included in the study.ResultsOf 2567 new sputum smear-negative TB patients, 1973 (90%) had sputum specimens examined at the end of the intensive phase, of whom 36 (2%) were smear-positive: the majority (n = 28) successfully completed treatment with either the same or a re-treatment regimen. At treatment completion, 1766 (85%) patients had sputum specimens examined, of whom 16 (0.9%) were smear-positive: all these were changed to a re-treatment regimen. Amongst the sputum-positive patients identified as a result of follow up (n = 52), four were diagnosed with multi-drug resistant TB (MDR-TB), three of whom were detected after smear examination at the end of treatment.ConclusionsGiven the high burden of TB in India, a 0.9% additional yield of smear-positive sputum smears at the end of treatment translates to 3,297 cases of smear-positive PTB. End-of-treatment smear is a low-yield strategy for detection of smear-positive TB cases, although further studies are needed to determine its population-level impact and cost, particularly in relation to other TB control interventions.

Rapid diagnosis of malaria using plasmodium lactate dehydrogenase (OptiMAL) test

Abstract We compared the sensitivity and specificity of OptiMAL test and Quantitative Buffy Coat(QBC) method with both thick & thin peripheral blood smear microscopic examination (gold standard) for diagnosing malaria in 103 children aged 1–15 years admitted with clinical suspicion of malaria. Of 103 cases, PBS study was positive in 59 (57.3%) children, QBC was positive in 60 (58.2%) and OptiMAL test was positive in 59 (57.3%) cases. Sensitivity and specificity of OptiMAL test was 100% and 97.8% while sensitivity and specificity of QBC method was 100% and 91.3%, respectively. OptiMAL test is a simple, sensitive and specific test for timely diagnosis of malaria in children.

Relationship between oxidative stress and clinical–pathological aspects in dogs experimentally infected with Rangelia vitalii

The aim of this study was to evaluate lipid peroxidation, protein oxidation and activity of enzymes that are indicators of oxidative stress in Rangelia vitalii infection in dogs. Animals were divided into two groups: negative control (n=5) and infected with R. vitalii (n=7). After inoculation, the parasitemia was estimated daily by microscopic examination of smears. Lipid peroxidation (TBARS) and advanced oxidation protein products (AOPP); and delta-aminolevulinate dehydratase (δ-ALA-D), superoxide dismutase (SOD) and catalase (CAT) activities in blood were evaluated. The samples were collected at days 10 and 20 post-inoculation (PI). TBARS and AOPP levels were higher in the infected group in both analyzed periods (P<0.01). The δ-ALA-D activity was reduced in blood of dogs infected with R. vitalii on days 10 and 20 PI. SOD activity was significantly increased (P<0.01) in the blood of dogs infected with R. vitalii at days 10 and 20 PI, while CAT activity was significantly increased (P<0.01) only at day 20 PI when compared to non-infected animals. A positive correlation was observed between the degree of parasitemia and TBARS and AOPP levels and activity of antioxidant enzymes. The δ-ALA-D activity was negatively correlated with the degree of parasitemia. Based on the increased levels of TBARS, AOPP, SOD and CAT activities, and inhibition δ-ALA-D activity, we concluded that dogs experimentally infected with R. vitalii develop a state of redox unbalance and that these changes might be involved in the pathophysiology of disease.

Age-Stratified Bayesian Analysis To Estimate Sensitivity and Specificity of Four Diagnostic Tests for Detection of Cryptosporidium Oocysts in Neonatal Calves

There is no gold standard diagnostic test for the detection of bovine cryptosporidiosis. Infection is usually highest in 2-week-old calves, and these calves also excrete high numbers of oocysts. These factors may give rise to variations in the sensitivity and specificity of the various diagnostic tests used to detect infection in calves of various ages. An age-stratified Bayesian analysis was carried out to determine the optimum diagnostic test to identify asymptomatic and clinical Cryptosporidium sp. infection in neonatal calves. Fecal samples collected from 82 calves at 1 week, 2 weeks, 3 weeks, and 4 weeks of age were subjected to the following tests: microscopic examination of smears stained with either phenol-auramine O or fluorescein isothiocyanate (FITC)-conjugated anti-Cryptosporidium monoclonal antibody, nested-PCR, and quantitative real-time PCR. The results confirmed a high prevalence of Cryptosporidium sp. infection, as well as a high level of oocyst excretion, in 2-week-old calves. The sensitivities of all the tests varied with the age of the calves. Quantitative real-time PCR proved to be the most sensitive and specific test for detecting infection irrespective of the age of the calf. The microscopic techniques were the least sensitive and exhibited only moderate efficiency with 2-week-old calves excreting large numbers of oocysts, the majority of which were diarrheic. It was concluded that, when interpreting the results of routine tests for bovine cryptosporidiosis, cognizance should be taken of the sensitivity of the tests in relation to the age of the calves and stage of infection.

An unusual febrile nonhemolytic reaction occurred after transfusion in a thalassemia major patient with asymptomatic Plasmodium falciparum infection

Febrile nonhemolytic transfusion reactions occur in 0.12% of transfusions, usually during transfusion or within 4 to 6 hours after transfusion and are not medically dangerous. A patient with thalassemia from Togo with asymptomatic malaria in which the infection became clinically manifest only after blood transfusion, mimicking a febrile nonhemolytic transfusion reaction, is presented. Thirty-two hours after transfusion of 2 O D- red blood cell (RBC) units, the patient (phenotype A(2) D+) developed fever and multiorgan failure and was admitted to the intensive care unit. Direct and indirect antiglobulin tests were negative on posttransfusion samples. Blood cultures and infectious diseases testing were negative. No malaria parasites were found at thick blood smear microscopic examination on Days 1 and 2 and the malaria rapid diagnostic test gave inconsistent results. Plasmodium total antibodies were detected in the serum at high levels. On Day 5, routine microscopic examination of blood smear revealed the presence of parasites in a very small number of RBCs. This finding was almost simultaneous to the availability of polymerase chain reaction testing results that were positive for P. falciparum. The sequential agglutination with anti-A antiserum allowed patient's and donors' RBCs to be separated and revealed that the parasitized cells were almost exclusively those of donors (14.4% vs. 0.029%). Malaria infection in implicated donors was excluded. In this patient with thalassemia with asymptomatic malaria, the infusion of two normal RBC units provided a favorable environment for a rapid parasite replication leading to a dramatic acute malaria attack.

Subtype analysis of Blastocystis isolates from symptomatic patients in Egypt

Blastocystis sp. has been described as the most common intestinal parasite in humans and has an increased impact in public health. To improve our understanding of the molecular epidemiology of this human-emerging parasite, we determined the Blastocystis subtypes (STs) and their relative frequency in Egyptian patients living in or in the vicinity of Cairo and presenting gastrointestinal symptoms. We obtained a total of 20 stool samples identified as positive for Blastocystis by microscopic examination of smears. Genotyping using partial small subunit ribosomal RNA gene analysis identified a total of 21 Blastocystis isolates corresponding to 19 single infections and one mixed infection (ST1 and ST3). Three STs were identified: ST3 was the most common ST in the present Egyptian population (61.90%) followed by ST1 (19.05%) and ST2 (19.05%). Together with previous studies carried out in different areas in Egypt, a total of five STs (ST1, ST2, ST3, ST4, and ST6) have been found in symptomatic patients. These data were compared to those available in the literature, and we underlined variations observed in the number and relative proportions of STs between and within countries. On the whole, it seemed that Blastocystis infection is likely not associated with specific STs even if some STs are predominant in the epidemiologic studies, but rather with a conjunction of factors in the course of infection including environmental risk and parasite and host factors.

Comparison of different methods for delayed post-mortem diagnosis of falciparum malaria

BackgroundBetween 10,000 and 12,000 cases of imported malaria are notified in the European Union each year. Despite an excellent health care system, fatalities do occur. In case of advanced autolysis, the post-mortem diagnostic is impaired. Quicker diagnosis could be achieved by using rapid diagnostic malaria tests.MethodsIn order to evaluate different methods for the post-mortem diagnosis of Plasmodium falciparum malaria in non-immunes, a study was performed on the basis of forensic autopsies of corpses examined at variable intervals after death in five cases of fatal malaria (with an interval of four hours to five days), and in 20 cases of deaths unrelated to malaria. Detection of parasite DNA by PCR and an immunochromatographic test (ICT) based upon the detection of P. falciparum histidine-rich protein 2 (PfHRP2) were compared with the results of microscopic examination of smears from cadaveric blood, histopathological findings, and autopsy results.ResultsIn all cases of fatal malaria, post-mortem findings were unsuspicious for the final diagnosis, and autoptic investigations, including histopathology, were only performed because of additional information by police officers and neighbours. Macroscopic findings during autopsy were unspecific. Histopathology confirmed sequestration of erythrocytes and pigment in macrophages in most organs in four patients (not evaluable in one patient due to autolysis). Microscopy of cadaveric blood smears revealed remnants of intraerythrocytic parasites, and was compromised or impossible due to autolysis in two cases. PCR and ICT performed with cadaveric blood were positive in all malaria patients and negative in all controls.ConclusionIn non-immune fatalities with unclear anamnesis, ICT can be recommended as a sensitive and specific tool for post-mortem malaria diagnosis, which is easier and faster than microscopy, and also applicable when microscopic examination is impossible due to autolysis. PCR is more expensive and time-consuming, but may be used as confirmatory test. In highly endemic areas where asymptomatic parasitaemia is common, confirmation of the diagnosis of malaria as the cause of death has to rely on histopathological findings.

Lipid peroxidation in cats experimentally infected with Trypanosoma evansi

The objective of this study was to evaluate the lipid peroxidation and the susceptibility of erythrocytes to in vitro peroxidation as indicators of oxidative damage in erythrocytes and their roles in the pathogenesis of anemia during experimental Trypanosoma evansi infection in cats. Animals were divided into two groups: control and infected with T. evansi. Seven cats were infected with 10(8) trypomastigotes each, and parasitemia was estimated daily for 49 days by microscopic examination of smears. Hematological and biochemical parameters were evaluated for monitoring of the disease. Plasma lipid peroxidation (Thiobarbituric Acid Reactive Substances (TBARS)) and the susceptibility of erythrocytes to in vitro peroxidation were evaluated. Blood samples for analysis were collected at days 21 and 49 post-inoculation. TBARS level, indicated by MDA concentration, was higher in the infected group than in the control group in both analyzed periods, as well as the in vitro erythrocyte peroxidation (P < 0.001). The infected cats had variable degrees of regenerative anemia, which could be explained by the damage in erythrocyte membrane caused by lipid peroxidation.

Prevalence of Cryptosporidium isolated from dromedary camels (Camelus dromedarius) in Qeshm Island, Southern Iran

This study was undertaken to assess the prevalence of Cryptosporidium in dromedary camels (Camelus dromedarius) on Qeshm Island, southern Iran. Sixty-five faecal samples were obtained from camels (age range 2–12 years) housed in ten privately owned herds from three equal-length areas (west, east and central) on Qeshm Island, southern Iran. All samples were analysed for Cryptosporidium oocysts by microscopic examination of smears stained by modified Ziehl-Neelsen technique. The parasite was detected in 11 camels (16.9%). The genus identity of the oocysts was confirmed by morphology and was similar to Cryptosporidium parvum. The mean (±SD) size of 225 oocysts was 5.1 (0.42) × 4.38 (0.27) µm (range 4.2–5.3 × 3.8–4.5 µm) with a shape index (length/width) of 1.17 (0.03; range 1.11–1.18). Data analyses indicated significant difference in prevalence of Cryptosporidium infection in western and central areas compared with the eastern area. Numerous reports indicate that water is a major vehicle for transmission of cryptosporidiosis. Geographical studies in western and eastern areas show that areas of vegetation, lagoons and seasonal rivers in these two regions are more frequent than the eastern part. Possible source of contamination of these waters by Cryptosporidium may be related to sewage from human and faeces of domesticated and wild animals. Therefore, it is possible that water may be a predisposing factor for transmission of cryptosporidiosis in these areas.

Trypanosoma evansi: Levels of copper, iron and zinc in the bloodstream of infected cats

The aim of this study was to evaluate the concentrations of copper, iron and zinc in blood serum of cats experimentally infected with Trypanosoma evansi. Animals were divided into two groups: control and infected with T. evansi. The animals were infected with 10 8 trypomastigotes each and parasitemia was estimated daily for 56 days by microscopic examination of smears. Hematological and biochemical parameters were evaluated for monitoring of the disease. Serum metal levels were determined in blood samples collected at days 7, 28 and 56 of the experiment. Inductively coupled plasma optical emission spectrometry was used to measure the levels of copper, iron and zinc. Significant differences were observed among groups ( P < 0.05). Increased levels of copper and decreased iron and zinc levels were observed. A decrease in the number of red blood cells was also observed 7 days after inoculation. Biochemical parameters were not altered. Therefore, the infection by T. evansi might alter the serum metal levels, causing metabolic disturbances in cats.

PCR diagnosis of visceral leishmaniasis in an endemic region, Mymensingh district, Bangladesh

Detection of Leishmania parasites in a clinical sample is necessary to confirm a suspected case of leishmaniasis. We compared the sensitivity of internal transcribed spacer 1-PCR (ITS 1-PCR) assay for parasite diagnosis with that of microscopic detection in clinical samples from kala-azar (KA) or post-kala-azar dermal leishmaniasis (PKDL) suspects in Mymensingh. Of 39 specimens collected from 35 KA and four PKDL suspects, 26 were positive by microscopic examination of smears from bone marrow and skin exudates; 38 specimens spotted on filter paper and 27 of the 28 Giemsa-stained slides tested by PCR proved positive by ITS1-PCR.

Molecular epidemiology of human Blastocystis isolates in France

Blastocystis sp. is the most common eukaryotic parasite in the intestinal tract of humans. Due to its strong impact in public health, in this study, we determined the frequency of different Blastocystis subtypes in patients in France. We hypothesized on the mode of transmission and tested a possible relationship between the subtype and symptomatic status. We obtained a total of 40 stool samples identified as positive for Blastocystis by microscopic examination of smears. Participants consisted of 25 symptomatic and 15 asymptomatic patients, for whom clinical and parasitological data were collected. For nested-polymerase chain reaction and genotyping, DNA was extracted directly from fecal samples or from fecal cultures. Morphological forms observed in fecal cultures were uncorrelated with symptomatic status. Genotyping using partial small subunit rRNA gene analysis identified a total of 43 Blastocystis isolates corresponding to 37 single infections and three mixed infections by two different subtypes. These 43 isolates belonged to five subtypes (1, 2, 3, 4, and 7) with predominance of subtype 3 (53.5%). Patient symptomatic status was uncorrelated with Blastocystis subtype.

Investigative modalities in infectious keratitis

Standard recommended guidelines for diagnosis of infectious keratitis do exist. Based on an extensive Medline literature search, the various investigative modalities available for aiding the diagnosis of microbial keratitis have been reviewed and described briefly. Preferred practice patterns have been outlined and the importance of routine pre-treatment cultures in the primary management of infectious keratitis has been highlighted. Corneal scraping, tear samples and corneal biopsy are few of the specimens needed to carry out the investigative procedures for diagnosis and for initiating therapy in cases of microbial keratitis. In bacterial, fungal and amoebic keratitis, microscopic examination of smears is essential for rapid diagnosis. Potassium hydroxide (KOH) wet mount, Gram's stain and Giemsa stain are widely used and are important for clinicians to start empirical therapy before microbial culture results are available. The usefulness of performing corneal cultures in all cases of suspected infectious keratitis has been well established. In cases of suspected viral keratitis, therapy can be initiated on clinical judgment alone. If a viral culture is needed, scrapings should directly be inoculated into the viral transport media. In vivo confocal microscopy is a useful adjunct to slit lamp bio-microscopy for supplementing diagnosis in most cases and establishing early diagnosis in many cases of non-responding fungal and amoebic keratitis. This is a non-invasive, high resolution technique which allows rapid detection of Acanthamoeba cysts and trophozoites and fungal hyphae in the cornea long before laboratory cultures give conclusive results. Other new modalities for detection of microbial keratitis include molecular diagnostic techniques like polymerase chain reaction, and genetic finger printing by pulsed field gel electrophoresis.

Prevalence ofCryptosporidiumspp. (Eucoccidiorida: Cryptosporiidae) in seven species of farm animals in Tunisia

1,001 faecal samples were obtained from 89 sheep (lambs and adult), 184 goats, 190 horses, 178 rabbits, 110 camels, 200 broiler chicken and 50 turkeys housed in farms from different localities in Tunisia. All samples were analysed for Cryptosporidium oocysts by microscopic examination of smears stained by modified Ziehl Neelsen technique. The parasite was detected in ten lambs and adult sheep (11.2 %) and nine broiler chicken (4.5 %). Molecular characterization, performed in four animals, identified C. bovis in three lambs and C. meleagridis in one broiler chicken. This work is the first report on Cryptosporidium in farm animals in Tunisia.