Review on diagnosis and molecular characterization of Toxoplasma gondii in humans and animals.

Toxoplasma gondii is an obligate intracellular protozoon which causes toxoplasmosis, an important zoonotic disease that is endemic worldwide. Common sources of T. gondii infection in humans are food or water contaminated with oocysts and raw or undercooked meat with cysts. In animals, common sources of infection include feed, water, or litter contaminated with oocysts. The diagnosis and molecular characterization of T. gondii infection in humans and animals is crucial due to public and veterinary health importance. Various traditional and serological methods have been used in clinical practice for toxoplasmosis diagnosis, but interpreting the results remains a challenge. Several molecular techniques have also been used for the detection and genetic characterization of T. gondii , but primarily in research settings. In this paper, we review the techniques that are currently used for the diagnosis and genetic characterization of T. gondii in humans and animals, along with their advantages and disadvantages. The techniques reviewed have laid the groundwork for the future development of more effective and precise detection and characterization of T. gondii . These advances will contribute to a better understanding of epidemiology, prevention and control of toxoplasmosis. Thus, this review would be of particular interest to clinical physicians, veterinarians and researchers.

Review on diagnosis and molecular characterization of Toxoplasma gondii in humans and animals

@#Toxoplasma gondii is an obligate intracellular protozoon which causes toxoplasmosis, an important zoonotic disease that is endemic worldwide. Common sources of T. gondii infection in humans are food or water contaminated with oocysts and raw or undercooked meat with cysts. In animals, common sources of infection include feed, water, or litter contaminated with oocysts. The diagnosis and molecular characterization of T. gondii infection in humans and animals is crucial due to public and veterinary health importance. Various traditional and serological methods have been used in clinical practice for toxoplasmosis diagnosis, but interpreting the results remains a challenge. Several molecular techniques have also been used for the detection and genetic characterization of T. gondii, but primarily in research settings. In this paper, we review the techniques that are currently used for the diagnosis and genetic characterization of T. gondii in humans and animals, along with their advantages and disadvantages. The techniques reviewed have laid the groundwork for the future development of more effective and precise detection and characterization of T. gondii. These advances will contribute to a better understanding of epidemiology, prevention and control of toxoplasmosis. Thus, this review would be of particular interest to clinical physicians, veterinarians and researchers.

Human MAIT Cell Activation In Vitro.

Mucosal-associated invariant T (MAIT) cells are an abundant innate-like T cell subset in humans, enriched in mucosal tissues and the liver. MAIT cells express a semi-invariant T cell receptor (TCR) and recognize microbial-derived riboflavin metabolites presented on the MHC Class I-like molecule MR1. In addition to activation via the TCR, MAIT cells can also be activated in response to cytokines such as IL-12 and IL-18, in contrast to conventional T cells. Here we describe TCR-dependent and -independent methods for MAIT cell activation. The TCR-dependent approaches include stimulation with microbead- or plate-bound anti-CD3/anti-CD28 antibodies, and with 5-OP-RU or paraformaldehyde (PFA)-fixed E. coli in the presence of antigen-presenting cells (APCs). The latter method includes a combination of TCR- and cytokine-mediated stimulation. The TCR-independent methods include direct stimulation with the recombinant cytokines IL-12 and IL-18, and indirect stimulation with TLR-4/TLR-8 agonists or influenza A virus in the presence of APCs. Finally, we outline a protocol to analyze activated MAIT cells using flow cytometry.

Bacteriology and antibiotic sensitivity patterns of urinary tract infections in a tertiary hospital in Bangladesh.

Urinary tract infection (UTI) is common bacterial infection in mankind. The changing antimicrobial sensitivity in UTI demands use of appropriate antibiotics. This prospective study was performed in Comilla Medical Collage during the period of January 2011 to December 2011. Five hundred and fifty one urine specimens from clinically suspected UTI patients were examined by Semi quantitative culture method and their antimicrobial sensitivity patterns were determined by disc diffusion technique. The study was designed to isolate and identify the nature of bacteria in UTI with their sensitivity pattern to common antibiotics. Of the 551 tested sample 131 samples showed growth of pathogens among which the most prevalent were E. coli 98(75%) followed by Klebsiella pneumonia 14(10.7%) and Enterococcus 8(6%). The majority 96(73.3%) of the isolates were from female while the remaining were from male and this sex difference was statistically significant (p<0.001). Imipenem, meropenem, amikacin, nitrofurantoin, gentamycin, mecillinum and amoxyclav are found to be effective against 76-100% of the uropathogens. Most powerful antibiotics in our study were imipenem, meropenem, amikacin and nitrofurantoin which show their efficacy against 91-100% isolates. In more than 60% case shows their resistance against amoxycillin, nalidexic acid, cefixime, ciprofloxacin, cotrimoxazole and cephalosporins which raises the question regarding rationality to empirically use of these antibiotics in UTI with out culture and sensitivity reports.

ROS-PIASγ cross talk channelizes ATM signaling from resistance to apoptosis during chemosensitization of resistant tumors.

With the existing knowledge of ATM's role in therapeutic resistance, the present study aimed at identifying the molecular mechanisms that influence ATM to oscillate between chemoresistance and chemosensitivity. We observed that the redox status of tumors functions as a major determinant of ATM-dependent 'resistance-to-apoptosis' molecular switch. At a low reactive oxygen species (ROS) condition during genotoxic insult, the ATM/sumoylated-IKKγ interaction induced NFκB activation that resisted JNK-mediated apoptosis, whereas increasing cellular ROS restored ATM/JNK apoptotic signaling. A search for the upstream missing link revealed that high ROS induces oxidation and ubiquitin-mediated degradation of PIASγ, thereby disrupting PIASγ-IKKγ cross talk, a pre-requisite for IKKγ sumoylation and subsequent NFκB activation. Interruption in the PIASγ-mediated resistance pathway channels ATM signaling toward ATM/JNK pro-death circuitry. These in vitro results also translated to sensitive and resistant tumor allograft mouse models in which low ROS-induced resistance was over-ruled in PIASγ knockout tumors, while its overexpression inhibited high ROS-dependent apoptotic cues. Cumulatively, our findings identified an unappreciated yet critical combinatorial function of cellular ROS and PIASγ in regulating ATM-mediated chemosensitization of resistant tumors. Thus, therapeutic strategies employing ROS upregulation to inhibit PIASγ during genotoxic therapy may, in future, help to eliminate the problems of NFκB-mediated tumor drug resistance.

Hypersensitivity reaction to all drugs of category-1 anti-tuberculosis regime in an adult tuberculosis patient.

Adverse drug reaction to tuberculous chemotherapy is not an uncommon problem. Usually it occurs to single drug and can be treated easily with minimal intervention. We follow WHO recommended guideline for National Tuberculosis Control Programs to treat these adverse reactions. Here we found an adult who has been suffering left sided pleural tuberculosis developed anaphylactic reaction to first dose of category-1 anti-TB regime. Later on it was found that he could not even tolerate smaller challenging doses of isoniazide, Ethambutol, Rifampicin and Pyrazinamide separately. It became very difficult to choose an alternate regime for this patient. Lastly a regime with levofloxacin, streptomycin and clarithromycin was give to treat him and patient was recovered with this regime successfully. This experience will help in management of unusual drug reactions to anti-tuberculosis drugs.

Pulmonary metastases of a papillary adeno-carcinoma of ovary in a pregnant lady.

A 28 years young lady admitted to National Institute of Diseases of Chest and Hospital (NIDCH), Dhaka with the complaints of respiratory distress and cough for 2 months. She was in her last trimester of first pregnancy when she becomes dyspneic on exertion and at night. Gradually it increases in time and she become unable to walk when she consulted with specialists in Barisal. She was examined clinically and radiographically and found some opacity on both lung fields. She was on several antibiotics, but got no improvement. In the meantime, an emergency caesarean section was done to have a healthy baby and patient came to NIDCH for this unexplained breathlessness and cough. She was severely dyspneic, tachypnoic, cyanosed. There were bilateral coarse crepitations, bilateral opacities predominantly on both lower and middle zones of both lungs obscuring costo-phrenic angles on chest X-rays. CT scan of chest shows bilateral reticulo-nodular shadows with a homogenous opacity on right lower lung which enhances after contrast scanning. A CT guided FNAC was done from that opacity which revealed a papillary adeno-carcinoma with psammoma bodies on cytopathological study. Thereafter, to explore the primary site, thyroid gland, abdominal organs was assessed adequately and only positive finding was raised CA-125 (706 IU/ml) which was consistent with ovarian cancer. In this way, a primary ovarian carcinoma in a pregnant young lady with normal sized ovary that metastasizes to lung causing bilateral pleural effusion and lymphangitis carcinomatosa was explored. This was an unusual presentation of ovarian papillary adeno-carcinoma with cough and breathlessness at the last trimester of pregnancy in absence of any abdominal mass.

Restoration of tumor suppressor p53 by differentially regulating pro- and anti-p53 networks in HPV-18-infected cervical cancer cells.

Abrogation of functional p53 is responsible for malignant cell transformation and maintenance of human papilloma virus (HPV)-infected cancer cells. Restoration of p53 has, therefore, been regarded as an important strategy for molecular intervention of HPV-associated malignancies. Here we report that differential regulation of pro- and anti-p53 setups not only upregulates p53 transcription but also stabilizes and activates p53 protein to ensure p53-induced apoptosis in HPV-18-infected cervical cancer. Functional restoration of p53 can be achieved by non-steroidal anti-inflammatory drug celecoxib via multiple molecular mechanisms: (i) inhibition of p53 degradation by suppressing viral oncoprotein E6 expression, (ii) promoting p53 transcription by downmodulating cycloxygenase-2 (Cox-2) and simultaneously retrieving p53 from Cox-2 association and (iii) activation of p53 via ataxia telangiectasia mutated-/p38 mitogen-activated protein kinase-mediated phosphorylations at serine-15/-46 residues. That restored p53 is functional has been confirmed by its ability of transactivating Bax and p53-upregulated modulator of apoptosis, which in turn switch on the apoptotic machinery in these cells. Studies undertaken in biopsy samples of cervical carcinoma further validated celecoxib effect. Our approaches involving gene manipulation and pharmacological interference finally highlight that celecoxib alters pro- and anti-p53 networks, not in isolation but in concert, to rejuvenate p53-dependent apoptotic program in HPV-infected cervical cancer cells.

Value of mental health first aid training of advisory and extension agents in supporting farmers in rural Queensland.

INTRODUCTION: This study was a pilot project responding to the increasing levels of stress, depression and other mental health issues in Australian rural areas resulting from prolonged drought and a changing economic and social environment. METHODS: Thirty-two Advisory and Extension Agents (AEAs) attended a training course held in 2007 and 2008 in Queensland, Australia. A year after the training, data was collected to determine its value. Interviews were conducted with course participants and their supervisors and focus groups were held with stakeholders (farmers, agency staff and health professionals). RESULTS: The findings show that Mental Health First Aid training improved the participants' confidence level and their knowledge of mental health issues and increased their empathy toward persons with mental health problems. Furthermore, providing training on mental health issues to AEAs was perceived by stakeholders to be beneficial to both farmers and AEAs. CONCLUSION: This study demonstrated that stakeholders and course participants see this type of training as very much needed and highly beneficial. Further, providing training in mental health issues to rural service providers can be very beneficial to their farmer clients and their social network.

Inter- and intralaboratory comparison of Ehrlichia equi and human granulocytic ehrlichiosis (HGE) agent strains for serodiagnosis of HGE by the immunofluorescent-antibody test.

Human granulocytic ehrlichiosis (HGE) is usually diagnosed by immunofluorescent antibody (IFA) serology with Ehrlichia equi-infected neutrophils or HGE agent-infected cultured HL60 cells. The HGE agent and E. equi are antigenically diverse, and interpretation of serologic results is also often variable. Thus, we investigated the sensitivity and specificity of various HGE agent and E. equi antigens used for IFA diagnosis by three different laboratories. Serum samples from 28 patients with well-characterized HGE and 9 patients with suspected HGE who were investigated by PCR, blood smear examinations, and serology were used, along with 9 serum samples from patients with other rickettsial and ehrlichial infections. Each serum sample was tested with up to 10 different antigen preparations. Overall, qualitative IFA results agreed in 70% of the samples. Titers among antigens were similar (r = 0.89 to 0. 96), but titers of individual samples varied by fourfold or more in 5 of 81 (6%) of the serum samples. Sensitivity ranged from 100% to 82%, and specificity varied from 100% to 67%, but these differences were not significant, even among those tested in the same laboratory or between two different laboratories. Antibodies were detected in 14 to 44% of acute-phase sera from confirmed HGE patients. Most false-positive reactions resulted with Ehrlichia chaffeensis; when these sera were excluded, the specificity of most antigens was 91 to 100%. These data indicate that IFA results often agree and that IFA is useful for diagnosis of HGE in convalescence. However, without further standardization, variability among serologic tests using E. equi and HGE agent isolates for diagnosis of HGE will occasionally provide discrepant results and confound diagnosis.

Clinical and laboratory evolution of a culture-confirmed case of human granulocytic ehrlichiosis.

A 74-year-old man from suburban New York City, who was hospitalized because of chest pain and fever, was diagnosed as having human granulocytic ehrlichiosis on the eighth hospital day. Although leukocyte and platelet counts were normal on admission, they fell to abnormally low values then normalized prior to specific therapy against the human granulocytic ehrlichiosis agent. Intracytoplasmic inclusions suggestive of Ehrlichia were observed in up to six percent of granulocytes, and the human granulocytic ehrlichiosis bacterium was cultured in an HL 60 human promyelocytic cell line. The patient improved dramatically within 24 hours of doxycycline treatment, after failing to improve on various beta lactam antimicrobial agents. He was discharged from the hospital 14 days after admission. Because human granulocytic ehrlichiosis was not diagnosed until his eight hospital day, clinical and laboratory parameters prior to specific treatment were available. This case illustrates the clinical and laboratory evolution of the infection with human granulocytic ehrlichiosis agent in humans.

Biphenotypic (mixed myeloid/T-cell) extramedullary myeloid cell tumor.

A 32-year-old male with a 4-year history of chronic myelogenous leukemia (CML) in chronic phase for 4 years, then myeloid blast crisis for 7 months, developed diffuse bulky lymphadenopathy in association with a white blood count (WBC) of 17,100/mm3 with 70% blasts. Biopsy of a cervical lymph node revealed a blastic extramedullary myeloid cell tumor, which showed a biphenotypic (mixed myeloid/T-cell) immunophenotype. Chromosomal analysis revealed karyotypic features of both myeloid and lymphoid lineages. Although extramedullary myeloid cell tumor (EMT, granulocytic sarcoma, chloroma) is well known to occur in chronic myelogenous leukemia (CML), to our knowledge this is the first description of evolution of CML into a biphenotypic EMT.

Dose-related sevoflurane metabolism to inorganic fluoride in rabbits.

Serum concentrations and urinary excretion of inorganic fluoride (fluoride ion), a metabolite of sevoflurane, were measured by an ion-chromatographic analyzer after inhalation of three different concentrations of sevoflurane in adult, male Japanese white rabbits weighing 2.6-3.6 kg. Sevoflurane was administered at concentrations of 0% (control), 1%, 2% and 3% (Groups I, II, III and IV, respectively) through a sevoflurane vaporizer for 2 hr under controlled ventilation. Blood and urine samples were collected during and after termination of sevoflurane inhalation at scheduled time intervals for 24 hr. The total volume of urine, the urinary pH and the osmolality of serum and urine were not significantly different among any of the groups. Osmolality of the serum and urine was within normal range in all groups of animals. The mean serum peak values of fluoride ion were 0.7 +/- 0.5, 22.8 +/- 8.7, 31.8 +/- 11.0 and 41.5 +/- 13.2 microM (mean +/- SD) in groups I, II, III and IV, respectively. Peak values were recorded within 15 min after the termination of inhalation. The cumulative amounts of fluoride ion excreted in urine in 24 hr were calculated to be 5.0 +/- 1.6, 26.1 +/- 6.7, 41.4 +/- 11.3 and 64.3 +/- 18.0 mumol (mean +/- SD) in groups I, II, III and IV, respectively. Regression analysis revealed significant correlations between the formation and excretion of fluoride ion, and the dose of sevoflurane (r = 0.85, p less than 0.05 and r = 0.89, p less than 0.05, respectively). The authors conclude that the formation and excretion of fluoride ion after sevoflurane anesthesia is dependent on the dose of the drug.(ABSTRACT TRUNCATED AT 250 WORDS)

Determination by ion-exchange chromatography of trifluoroacetic acid as a biliary metabolite of isoflurane in the rabbit.

The optimal composition of eluant for trifluoroacetic acid (TFAA) analysis by ion-exchange chromatography was determined and a preliminary study to detect TFAA, a metabolite of isoflurane, in bile of rabbits during and after inhalation of isoflurane was made. The optimal composition of eluant for TFAA analysis by ion-exchange chromatography was determined to be a solution of 2mM Na2CO3 and 4mM NaHCO3. Rabbits were divided into 4 groups; A group: 1 hr inhalation of 2% isoflurane, B group: 2 hr inhalation of 2% isoflurane, C group: 2 hr inhalation of 3% isoflurane, and D group: 2 hr inhalation of 4% isoflurane. The maximum biliary TFAA concentration in groups A, B, C and D averaged 17.5 +/- 1.4 microM (mean +/- S.D.) at 105 min, 37.1 +/- 11.5 microM at 315 min, 54.5 +/- 9.4 microM at 585 min and 34.4 +/- 12.1 microM at 555 min after termination of inhalation. Cumulative excreted amounts of biliary TFAA for 20 hours were 746 nmols in A group (8 hours), 3,421 nmols in B group, 6,582 nmols in C group and 3,267 nmols in D group. Half-lives of biliary excreted TFAA were 63 min, 325 min, 478 min and 546 min, respectively. These results showed that ion-exchange chromatography is more convenient for detecting TFAA as a metabolite of isoflurane than isotachophoresis, gas chromatography, paper chromatography and thin layer chromatography and that TFAA was determined as a biliary metabolite of isoflurane in the rabbit.

Estimation of births averted in rural Bangladesh: a case study of Uthali.

PIP: The government of Bangladesh considers the population problem to be the major problem for the country. For the purpose of economic planners, the estimation of births averted under a program is an important aspect of any evaluation. The estimation of births averted is a measure of the quantitative change in the expectations, assessed in terms of probabilities, of future births to a cohort of women resulting from the adoption or modification of birth control practice by them or their husbands. The data used represent a cross section of women in rural society. The age specific continuation rates are calculated by single decrement life table technics. After calculating the estimates of births averted by the sample of acceptors, these numbers are converted to a ratio of births averted/1000 acceptors. The births averted/1000 women are 70 after 3 months, 142 after 6 months, 172 after 9 months, and 218 after 12 months. Findings relate only to use of oral contraception; additional births are averted by acceptors who, upon discontinuing their method for 1 reason or another, switch to another method. Based on the number of live births averted, the Uthali Family Planning Program will make an important contribution to fertility decline in Bangladesh's rural areas in the following decades.^ieng