Ziehl-Neelsen Research Articles

Криптоспоридиоз телят в условиях частного подворья на территории разных районов Карачаево-Черкесской республики

Cryptosporidiosis is a widespread parasitic (protozoal) disease. to which more than 170 species of animals are susceptible.
 Purpose of the study. To study the spread of cryptosporidiosis of calves in the conditions of private farmsteads in some areas on the territory of the Karachay-Cherkess Republic.
 Materials and methods. Fecal samples from 55 calves (clinically healthy and sick, with signs of diarrhea) aged 3 to 50 days were examined for the presence of the causative agent of cryptosporidiosis using the native smear method. The finished smears were fixed according to the Nikiforov method, stained with carbolic fuchsin according to Ziehl–Neelsen and examined under a binocular light microscope «Micromed-3» with an immersion lens. Cryptosporidium oocysts acquire a rich pink-red color when stained by Ziehl–Neelsen. The final diagnosis was established based on the detection of oocysts characteristic of this invasion. Cryptosporidium was identified using the «Determinant of parasitic protozoa» of Krylov M.V. and the atlas of Vershinin I.I.
 Results of the study. Microscopy of smears of faecal samples from 55 calves revealed cryptosporidium oocysts in 29 smears, thus, the extent of invasion (EI) was 52.7 %.

Enteric Protozoan Parasitosis and Associated Factors among Patients with and without Diabetes Mellitus in a Teaching Hospital in Ghana.

Enteric protozoa infections (EPIs) could worsen clinical outcomes in patients with diabetes mellitus and therefore requires prompt and accurate diagnosis and attention. This study aimed to determine the burden of EPIs and their associated factors among patients with and without diabetics at the Komfo Anokye Teaching Hospital (KATH) in Ghana. Again, the diagnostic performance of parasitological techniques routinely used for diagnosis was assessed. A total of 240 participants (made up of 140 patients with diabetes and 100 patients without diabetes) were recruited into the study by simple random sampling from November 2020 to May 2021. Stool samples of participants were collected, along with their demographic information, and examined using the saline direct wet mount (DWM), formol-ether concentration (FEC), and modified Ziehl-Neelsen staining (ZNS) techniques for the presence of enteric protozoans. Enteric protozoa were found among 62/140 (44.3%) diabetic patients and 13/100 (13.0%) nondiabetic patients. The predominant protozoa identified were Cryptosporidium spp. (17.86%) among patients with diabetes and Blastocystis hominis (7.0%) among patients without diabetes. EPI was associated with diabetes mellitus status (AOR = 3.48, 95% CI, 1.55-7.79), having diabetes for more than five years (AOR = 3.83, 95% CI, 1.65-8.86) and having comorbidity (AOR = 2.93, 95% CI, 1.33-6.45). The FEC technique had the highest sensitivity (100.0%), specificity 94.3% (95% CI, 91.35-97.22), and accuracy 95.0% (95% CI, 88.54-98.13) when compared to other techniques for diagnosis. EPIs are a significant health problem among patients with diabetes at KATH, and therefore antiparasitic drugs should be included in their treatment protocols for better health outcomes. Again, the FEC technique has demonstrated better performance in detecting EPIs and is therefore recommended to achieve early and accurate diagnosis of EPIs.

Identification of Cryptosporidium parvum and Blastocystis hominis subtype ST3 in Cholga mussel and treated sewage: Preliminary evidence of fecal contamination in harvesting area

Cryptosporidium parvum and Blastocystis hominis are foodborne parasites known for causing diarrhea. They accumulate in mussels grown on contaminated water bodies, due to the discharge of treated sewage from sewage treatment plants (STP). Despite this, some countries like Chile do not include these parasites in the control or monitoring of sewage water. The objective of this research was to evaluate the contamination of C. parvum. and B. hominis from treated sewage (disinfected by chlorination) and Cholga mussels in a touristic rural cove from the bay of Concepción. Cholga mussels from commercial stores and a treated sewage sample were analyzed. Cryptosporidium spp. was identified by Ziehl-Neelsen-Staining (ZNS) and C. parvum by direct-immunofluorescence assay (IFA) from ZNS-positive samples. Blastocystis hominis was identified by PCR using locus SSU rDNA. C. parvum and B. hominis subtype ST3 were found in 40% and 45% of Cholga mussel samples, respectively, and both parasites were identified in the treated sewage. Blastocystis hominis SSU rDNA gene alignment from Cholga mussels and treated sewage showed 89% of similarity, indicating that could be the same parasite in both samples. We describe the first evidence of possible contamination with these parasites from treated sewage to Cholga mussel suggesting an environmental contamination with high human risk. Based on these results, further studies will consider all the rural coves and STP from the bay to prevent possible contamination of these parasites.

Disruption of a kasB homolog gene (kasB) causes attenuation of cell invasion and virulence of Nocardia seriolae.

Nocardia seriolae is the primary aetiological agent of nocardiosis in fish, which causes mass mortality in freshwater and marine fish. β-ketoacyl-ACP synthase (KAS) is one of the essential enzymes in the synthesis of mycolic acids (MASs) in Mycobacterium spp. and has been chosen as the target for therapeutic intervention in mycobacterial diseases. In the present study, a kasB homologue gene (kasB) was identified in the genome of N. seriolae, and the gene-deficient mutant (ΔkasB) was generated based on a clinical isolate, XSYC-Ns. Compared to the wild-type (WT) strain, the ΔkasB showed a measurably growth defect invitro but retained the acid-fastness in acid-fast staining. Observation of the cell ultrastructure showed some alterations in the cell wall of the ΔkasB strain. Compared to its original strain, the cell wall lipid layer seemed sparser, and a wider electron-transparent zone was observed in the cell wall of ΔkasB strain. Moreover, the ΔkasB strain showed impaired ability of cell invasion as well as intracellular survival in the cell line originating from the head-kidney of the large yellow croaker (LYC-hK), compared to its original strain. In addition, the deficiency of ΔkasB significantly attenuated the virulence of N. seriolae in largemouth bass. The present study suggested that the ΔkasB gene might be involved in the synthesis of extracellular cell-wall lipids in N. seriolae and play a crucial role in its pathogenicity.

Copro-prevalence of <i>Cryptosporidium </i>in pigs of selected districtsin West coast of India: A preliminary study

In India, Cryptosporidium spp. have been detected and characterised from humans and domestic animals, mostly ruminants. Although pigs can act as an important reservoir of Cryptosporidium infection to humans, comprehensive studies have not been conducted in pig population in India. Hence, present study attempt to elucidate the prevalence and diversity of Cryptosporidium infection in pigs in the selected districts of three states situated in the West coast of India. The pig faecal samples (n=221) were subjected to coprological examination using modified Ziehl Neelsen (mZN) staining method, followed by polymerase chain reaction to detect Cryptosporidium spp. Using mZN staining method, 1.35% (95% CI, 0.46%–3.91%) of the samples were found positive for Cryptosporidium oocyst. However, the DNA from Cryptosporidium positive faecal samples could not be amplified using polymerase chain reaction probably because of the low sensitivity of the PCR and or low oocyst number in the faecal sample. Cryptosporidium spp. of pig origin have zoonotic potential and a certain proportion of pigs infected with Cryptosporidium may be apparently healthy. Therefore, the pig farmers need to be made aware of hygienic practices while handling pigs and pig manure and and general public must be made sensitised about the good agricultural practices and standard food hygiene practices to prevent foodborne Cryptosporidium infection.

Transbronchial brushing cytology and paired biopsy in endobronchial tuberculosis: A report of 72 cases focusing on the morphological features

Objectives: The objectives of this study were to review the transbronchial brushing cytology and histological specimens of endobronchial tuberculosis (EBTB) and to explore the morphological features, diagnostic pitfalls, and dilemmas. Material and Methods: Transbronchial brushing cytology and concurrent biopsy specimens obtained between July 2017 and June 2020 were reviewed. EBTB was confirmed based on the clinical response to the anti-TB treatment in addition to the positive findings of at least one of the following methods: Acid-fast bacilli stain (AFB), auramine-rhodamine stain (A-R), detection of TB bacterial DNA (TB-DNA) by polymerase chain reaction, T-cell spot test (T-spot), and typical pathologic changes of TB on cytology or bronchoscopy biopsy. A total of 72 confirmed cases were studied. Results: Of the 72 patients, 42/72 (58.3%) and 30/72 (41.7%) were female and male patients, respectively. Bronchoscopic findings revealed five subtypes of EBTB, including inflammation infiltration, ulceration necrosis, granulation hyperplasia, cicatrices stricture, and tracheobronchial malacia. AFB, A-R, TB-DNA, and T-spot were positive in 39, 26, 33, and 46 cases, respectively. The detection rate of necrosis in the cytological specimens (90.3%) was significantly higher than that in the biopsy specimens (77.8%; P < 0.01). The percentage of Langhans giant cells detected by cytology (13.9%) was significantly lower than that detected by the pathological examinations of the tissues (38.9%) (P < 0.01). The detection rates of metaplastic squamous cells and epithelioid cells showed no significant difference with respect to the cytology and biopsy findings. In addition to the two patients who had concurrent carcinomas, atypical cells were reported in nine patients through cytopathological diagnosis, among them two were suspected to have carcinomas, two were with the impression that spindle cell neoplasms could not be excluded, and the other five were considered as reactive atypia. Moreover, one biopsy could not rule out the well-differentiated squamous cell carcinoma. Conclusion: Some morphological variations may cause challenges in cytological evaluation. Moreover, diagnostic dilemmas can occur even in the assessments of tissue pathology.

Clinical profile of pulmonary tuberculosis patients in Oman from 2010 to 2014:A tertiary care hospital perspective

BackgroundDespite the efforts taken to improve the screening and treatment programs, Tuberculosis (TB) affects many people around the world. This descriptive retrospective study was conducted to explore the clinical profile of patients diagnosed with Pulmonary TB at a tertiary care hospital, Sultanate of Oman, during the period between 2010 and 2014. Methods.This is a cross-sectional study conducted in a tertiary care hospital in Sultanate of Oman during the period between 2010 and 2014. Clinical profile of pulmonary TB cases was investigated. The collected data included patient nationality, gender, age group, method of diagnosis, presenting symptoms, risk factors for TB and its complications. All data was gathered from electronic medical records. The data was analyzed by SPSS version 23. The study was ethically approved by the Ethical committee at the institute. ResultsOur study covered 239 patients with pulmonary and extra pulmonary TB. Pulmonary TB constituted 128 patients (54 %) of the total sample. More than half of the included patients were males and more than 70 % were Omanis. The mean age was 42.74±18.2 years. Half of the pulmonary TB patients fall within 25–50 years age range. Diabetes and hypertension were the most common risk factors reported at 24 patients (20 %) of the population each. While smoking was found to be one of the main risk factors to develop pulmonary TB, 23 patients (18 %) of the subject population were smokers and 13 patients (10 %) had a recent history of traveling along with 13 TB patients and 13 patients (10 %) had history of some kind of contact with TB patients. The main symptoms observed were cough in 86 patients (67 %), fever in 74 patients (58 %) associated with weight loss in 58 patients (45 %) and around 14 patients (11 %) were diagnosed incidentally. Out of the studied population, 22 patients were diagnosed with Ziehl–Neelsen (ZN) stain, followed by 25 patients with TB culture alone and 55 patients had both ZN and TB culture together. Pleural effusion and pneumothorax were the major complications experienced by the subject population, 13 (10.2 %) and 8 (6.3 %) patients respectively. However, 18 patients (14.1 %) died from pulmonary TB. ConclusionMost of the patients covered were of young to middle age, ranging between 25 and 50 years and most of them had pulmonary TB. Diabetes, hypertension, and smoking were found to be the main risk factors. The subject patients presented mainly suffered from cough, fever, and weight loss. ZN staining and sputum culture were the main methods of diagnosis. Pleural effusion and pneumothorax were the most common complications of pulmonary TB encountered in the subject population.

Parasit Gastrointestinal pada Kadal Naga Berjanggut (Pogona vitticeps) di Cijeruk, Bogor

Bearded dragons are reptiles that are quite popular as pets. There are many factors of disease that can infect bearded dragons. The environment and feed affect the level of disease threat, such as parasitic infection in the gastrointestinal tract. Data information related to parasitic gastrointestinal infections in bearded dragons in Indonesia is still limited, so supporting examinations are necessary to help diagnose. A total of 38 fecal samples were collected from the bearded dragon in Cijeruk Bogor. Samples were examined by simple flotation method, Ziehl Neelsen staining, and counting the number of parasitic eggs. All samples were positive for gastrointestinal parasite infections such as Choleoeimeria sp., Isospora amphiboluri, Cryptosporidium spp., ascarid egg, and oxyurid egg. The average number of Choleoeimeria sp. and Isospora amphiboluri oocysts were 8730 and 51.281 oocysts per gram of feces. The average number of helminth eggs per gram was 290 and 2325 for ascarid and oxyurid, respectively. Most of the infected bearded dragons do not show significant clinical symptoms. The number of oocysts and worm eggs per gram of feces showed mild and high results.

Breakthrough of chemiluminescence-based LAM urine test beyond HIV-positive individuals: Clinical diagnostic value of pulmonary tuberculosis in the general population.

To investigate the diagnostic value of a novel high-sensitivity urine lipoarabinomannan (LAM) test (chemiluminescence-based) for active tuberculosis in the general population. A retrospective study was conducted on 250 clinical suspected tuberculosis patients who were HIV-negative and visited the Fourth People's Hospital of Foshan from January 2022 to December 2022. Among them, there were 135 cases of pulmonary tuberculosis, 34 cases of extrapulmonary tuberculosis, and 81 cases of non-tuberculosis. Urine samples were collected for LAM antigen detection before treatment, and laboratory data of sputum smear acid-fast staining (smear method), sputum culture, and GeneXpert method were collected. Using clinical diagnosis as the reference standard, the diagnostic efficacy of 4 methods for detecting active tuberculosis was evaluated. For the 135 cases of pulmonary tuberculosis, the sensitivity of sputum smears, sputm culture, sputm GeneXpert method, and urine LAM were 29.6% (40/135), 45.9% (62/135), 59.3% (80/135), and 51.9% (70/135), respectively. The combination of LAM + GeneXpert and LAM + culture had the highest sensitivity for detecting active pulmonary tuberculosis, which were 71.0% and 78.2%, respectively. For the detection of sputum culture-negative pulmonary tuberculosis, the positive rates of smear, GeneXpert, and LAM were 0.0% (0/73), 53.4% (39/73), and 52.1% (38/73), respectively. LAM + smear and LAM + Genexpert could detect 52.1% and 68.5% of sputum culture-negative patients, respectively. The high-sensitivity urine LAM test holds promise for tuberculosis diagnosis in the general population. It demonstrates high-sensitivity, enabling the detection of sputum culture-negative pulmonary tuberculosis patients. Furthermore, when combined with existing methods, it can enhance the overall detection rate.

Prevalence and Molecular Identification of Cryptosporidium Species among Human Population in Zakho District, Duhok Province, Kurdistan Region, Iraq

This cross-sectional study was carried out to determine the prevalence and molecular identifications of Cryptosporidium species among the human population in Zakho district, Duhok Province, Kurdistan Region, Iraq. The study was performed during the period from August 2021 to March 2022 and involved 700 fecal specimens of both genders and different ages collected from Zakho General Hospital and pri¬mary schools located in the suburban area of Zakho City. The data were analyzed using statistical soft¬ware version SPSS (25). Cryptosporidium species oocyst was found in 87.57% (613/700) of the examined specimens using the Modified Ziehl Neelsen technique. Males showed a slightly higher infection rate than females (88.4% vs 85.5%) with a non-significant difference (P> 0.05). The highest infection rate (91.7%) was present in individuals between the ages of >30 to 40 years, with a statistically significant difference (P<0.05). Molecular identification of Cryptosporidium species was estimated using 18S-rRNA gene for Cryptosporidium genus using Nested PCR technique and genus-specific primers (Cry18 AS1 and Cry18 S2) that yielded 347 bps product indicating Cryptosporidium genus, then species-specific (Cry18 AS1 and Cry18 S1) were used for the detection of Cryptosporidium parvum and Cryptosporidium hominis. 45/50 samples were positive and revealed 240 bps. The acquired sequences were deposited into the GeneBank database under the accession numbers OP107010 for C. hominis, OP104444, OP153870, OP163188, OP163414, and OP164631 for C. parvum.

Detection and molecular characterization of Cryptosporidium spp. in dairy calves in Lisbon and Tagus Valley, Portugal

Cryptosporidium is a protozoan parasite with worldwide distribution, infecting a wide range of hosts with some zoonotic species. Calves have been identified as one of the most common reservoirs of this parasite. However, little is known about the genetics of Cryptosporidium in calves in Portugal. This study aimed to molecularly characterize infections of Cryptosporidium in pre-weaned calves from the Lisbon and Tagus Valley (LTV) in Portugal. Fifty-two samples were collected from calves from eight dairy and two beef farms in LTV, Portugal. Cryptosporidium oocysts were detected by Modified Ziehl-Neelsen staining (MZN) and direct immunofluorescent assay (DFA). MZN and DFA revealed the presence of Cryptosporidium oocysts in 40.4% (21/52) and 67.3% (35/52) samples, respectively. Positive samples were analyzed by PCR-RFLP of the 18 s rRNA gene for species identification. DNA amplification of the 18S rRNA gene was successful for 88.6% (31/35) of samples. Cryptosporidium parvum was identified in 96.8% (30/31) of the samples, and from one sample Cryptosporidium bovis was identified. Cryptosporidium parvum positive samples were subtyped by sequencing the PCR product of a partial fragment of the 60 kDa glycoprotein (gp60) gene. Subtype analysis of the C. parvum isolates revealed that all isolates belonged to subtype family IIa. Four subtypes were recognized within this subtype family, including the hyper-transmissible IIaA15G2R1 subtype that is the most frequently reported worldwide (27/30), IIaA14G2R1 (1/30), IIaA16G2R1 (1/30) and IIaA19G2R1 (1/30). To our knowledge, this is the first report of C. bovis, and C. parvum subtypes IIaA14G2R1 and IIaA19G2R1 in cattle in LTV, Portugal. The presence of the zoonotic C. parvum subtype in this study suggests that pre-weaned calves are likely to be a significant reservoir of zoonotic C. parvum, highlighting the importance of animal-to-human infection transmission risk. Further molecular studies are required to better understand the epidemiology of cryptosporidiosis in Portugal.

Pathomorphological characteristics of tuberculous placenta and its clinical implication

BackgroundThe study of pathologic diagnosis of placental TB is rare. The aim of this study is analyzing the pathomorphological characteristics of tuberculosis (TB) placenta during pregnancy and its clinical significance.MethodsNineteen cases of placental tissue specimens during pregnancy were collected from June 2015 to February 2022 at Shanghai Public Health Clinical Center, the only inpatient center for pregnant women with TB in Shanghai, China. Hematoxylin-eosin staining, acid-fast staining, and molecular testing were applied to analyze them comprehensively in combination with clinical information.ResultsAmong the 19 cases, 7 cases caused intrauterine stillbirth, 3 cases received artificial abortion required by the pregnant woman, the other 9 cases received standard delivery and the infants survived, however, 3 of them were low-weight preterm infants, and another 1 case suffered mild intrauterine asphyxia. The 9 surviving infants were followed-up, of which 3 cases got congenital TB. For pathological characteristics of placental tissues under light microscopy, there were 3 cases of epithelioid granuloma formation, 13 cases of acute fetal membranitis, 4 cases of caseous necrosis, 7 cases of inflammatory necrosis, 10 cases of coagulative necrosis, and 6 cases with small focal calcifications. All placental tissues were positive for acid-fast staining and polymerase chain reaction (PCR). Molecular pathological diagnosis showed that 18 cases were positive for Mycobacterium tuberculosis, with 1 case not having received examination.ConclusionsCombining acid-fast staining and molecular pathological testing is helpful for accurately diagnosing placental TB.

578. Performance of acid-fast stain, real-time PCR, and Xpert MTB/RIF in a bronchial washing for the Rapid Diagnosis of Mycobacterial infection in real-world

Abstract Background Mycobacterium culture remains still the gold standard for diagnosing mycobacterium infections, but there are some limitations. Acid-fast stain and simultaneous molecular methods are recommended for early diagnosis and treatment. Bronchial washing has been reported to be an effective diagnostic sample, even in cases sputum smear tests are negative. Additionally, it can be used in multiple assays. Therefore, we analyzed the clinical performance of acid-fast stain, real-time PCR, and Xpert MTB/RIF assay compared to mycobacterial cultures in the same bronchial washing. Methods We screened laboratory data from January 2018 to December 2020 at Jeonbuk National University Hospital. We enrolled the samples that all the tests for analysis, acid-fast stain, rPCR, Xpert MTB/RIF, and mycobacterial culture, were ordered simultaneously. The study was approved by the Institutional Review Board of Jeonbuk National University Hospital (CUH 2022-05-036-001). In short, the bronchial washing was sent to a clinical microbiology laboratory. The cultures were done with 2% Ogawa medium and BacT/Alert MB culture bottle after NALC-NaOH. The rPCR and the Xpert MTB/RIF was done according to the manufacturer's guide. The statistical analysis was done with Excel 2016 and Analyse-it. Results A total of 1966 samples were enrolled, and the culture-positive sample is defined when either the solid or liquid cultures showed positive. In the mycobacterium culture, 12.8% were MTB positive, and 10.7% were non-tuberculosis Mycobacterium (NTM) positive. The sensitivity and specificity of acid-fast stain were 45.6% and 98.9%, respectively, compared to mycobacterium culture positive, including MTB or NTM. The sensitivity and specificity of rPCR in MTB and NTM are 85.7%/94.3% and 52.9%/99.3%, respectively, and Xpert MTB/RIF in MTB cases was 94.8% and 92.1% (Table 1). Conclusion The acid-fast stain is still a valuable diagnostic tool despite its low sensitivity because it is easy to prepare and it doesn't require specialized equipment or facilities. The rPCR for MTB showed reliable results. However, in rPCR for NTM, the sensitivity was much lower than MTB. Xpert MTB/RIF showed superior sensitivity than rPCR here. The assays should be used strategically in suspected mycobacterium infections. Disclosures All Authors: No reported disclosures

Host factors associated with false negative results in an interferon‐γ release assay in adults with active tuberculosis

ObjectiveTo identify host factors associated with false-negative results of interferon-γ release tests in adults with active tuberculosis. MethodsThe clinical data of 943 patients with active tuberculosis diagnosed by acid-fast smear staining, Mycobacterium tuberculosis culture, Mycobacterium tuberculosis PCR and pathological examination at West China Hospital of Sichuan University were retrospectively analysed. According to the results of the interferon γ release test (IGRA), the patients were divided into the IGRA− group and IGRA+ group. Logistic regression was used to analyze the sociodemographic data and clinical characteristics of participants in the IGRA− group and IGRA+ group. ResultsAmong 943 patients with active tuberculosis, 174 (18.5 %) were IGRA negative (false negative), and 769 (81.5 %) were IGRA positive. Multivariate logistic regression analysis identified the following characteristics independently associated with IGRA negativity: age (OR: 1.02; 95 % CI: 1.01 1.03; p = 0.006), anti-tuberculosis treatment >1 month (OR: 1.68; 95 % CI: 1.12 2.52; p = 0.013), HIV infection (OR: 9.48; 95 % CI: 3.23 27.85; p = 0.000), combined with connective tissue diseases (OR: 2.78; 95 % CI: 1.30 5.94; p = 0.008) and low hemoglobin (OR: 0.99; 95 % CI: 0.98 1.00; p = 0.044) was associated with an increased false-negative probability of IGRA. ConclusionAge, anti-tuberculosis therapy >1 month, coinfection with HIV, coassociated connective tissue disease and decreased hemoglobin were identified as risk factors for false-negative results of IGRA. Our results suggest a careful interpretation of IGRA in adults with these characteristics.

A Study on the Bacteriological Profile of Ascitic Fluids and Their Antibiotic Susceptibility Pattern in a Tertiary Care Hospital.

Background Ascitic fluid culture remains an essential step in the management of all patients with ascites, regardless of their presenting complaints. Diagnostic paracentesis should not be delayed or prevent timely administration of antibiotics, particularly in unstable patients. Hence, it is an essential part of the surveillance system of every hospital to perform ascitic fluid culture and assess the antibiotic susceptibility patterns of bacterial isolates. In view of this perspective, the present study was conducted at Chengalpattu Medical College Hospital, Tamil Nadu, India. Objective The aim of the study isto determine the bacterial isolatesof ascitic fluid samples and study their antibiotic susceptibility patterns. Materials and methods Ascitic fluids received in the central laboratory at the Department of Microbiology from various departments were included in this study. Preliminary identification of isolates wasperformed by direct Gram staining, acid-fast staining, and motility testing by the hanging drop method. Within one hour of receiving the samples, they were plated onto blood agar and MacConkey agar media and incubated for 18-24 hours at 37°C for isolation. Growth was checked, and species identification was done based on conventional methods. Antibiotic susceptibility testing was performed using the Kirby-Bauer disk diffusion method. Results In this study, a total of 100 ascitic fluid samples were collected, of which only eight (8%) showed growth. Among the eight isolates, six (75%) were Gram-negative bacilli (GNB). Four (66.66%) of the six GNB wereKlebsiella spp., while the remaining two (33.33%) were Escherichia coli. Both Gram-positive cocci wereStaphylococcus aureus. All the GNB isolates were susceptible to meropenem, piperacillin-tazobactam, and ceftriaxone, with varying susceptibilities to other drugs. Both Gram-positive isolates were found to be methicillin-sensitive Staphylococcus aureus. Conclusion GNB were the predominant organisms in cases of ascitic fluid infection, and they showed 100% susceptibility to carbapenem drugs (especially meropenem),piperacillin-tazobactam, and ceftriaxone.All these drugs can be kept in reserve for serious infections. Amikacin and gentamicin showed promising susceptibility. These drugs can be started empirically with patients on admission before performing culture. Drug adjustments may be later made based on culture reports.

Microbiological persistence in patients with Mycobacterium abscessus complex lung disease: The prevalence, predictors, and the impact on progression

ObjectivesPersistent growth of Mycobacterium abscessus complex (MABC) in the respiratory system is not uncommon and may indicate continuous infection of MABC lung disease (MABC-LD), but its prevalence, risk factors, and clinical impact have not been investigated. MethodsThe present study was conducted in two medical centers in northern Taiwan. We enrolled patients with MABC-LD and investigated the prevalence and predictors of persistent culture positivity (MABC-PP). Furthermore, we analyzed the association between MABC-PP and radiographic or clinical progression. ResultsAmong 189 patients with MABC-LD, 58 were in the MABC-PP group. Independent predictors for MABC-PP included an increasing radiographic score and highest acid-fast stain (AFS) of strong positivity (3-4+) at initial diagnosis (compared with negative AFS). MABC-PP and highest AFS were independently associated with MABC-LD progression by the multivariable analysis model. The adjusted hazard ratio increased to 3.56 when the two independent factors existed. ConclusionsMABC-PP accounted for 30.7% and was predicted by initial AFS grade and radiographic score. Patients with MABC-PP, and highest AFS grade might have disease progression.

Molecular Detection of Mycobacterium avium paratuberculosis by Large Sequence Polymorphisms (LSPs)

Background: Mycobacterium avium subspecies paratuberculosis is a significant pathogen causing Johne’s disease. Rational evaluation of the prevalence of this organism in animals has been restricted by the lack of accurate diagnostic techniques. DNA microarray-based and computational study of Mycobacterium avium subsp. paratuberculosis strain K10 discovered 17 distinct large sequence polymorphisms present. Of these 17 LSPPs, 7 were with a specificity of greater than 98% and in some cases, sensitivity of up to 95% suggesting that the LSPs are ideal for diagnosing M. avium subsp. paratuberculosis. Methods: A total of 218 fecal samples were collected from cattle (n=180) and buffaloes (n=38) for detection of M. avium subsp. paratuberculosis (MAP) using Large Sequence Polymorphisms (LSPs) and IS900 PCR. The fecal samples were subjected to Ziehl-Neelsen staining, isolation and PCR. A total of 90 fecal samples were found ZN staining positive and were subjected to isolation of MAP, growths were seen in two fecal samples after 16 weeks of incubation. Result: Out of 218 fecal samples tested, 7 samples were positive by both LSP PCR viz; LSPP2 (MAP0284), LSPP4 (MAP0865), LSPP11(MAP2154) LSPP12 (MAP2182c), LSPP12 (MAP2188c), LSPP15 (MAP3774), LSPP16 (MAP3815) and IS900 sequence with an amplicon size of 600, 597, 375, 430, 719, 621, 611 and 229 bp. DNA was also extracted from the isolates (n=2) and were confirmed by PCR targeting Large Sequence Polymorphism (LSPs) and IS900. Detection of MAP in all the 7 fecal samples both by IS900 and by 7 LSPPs indicates the detection potential of selected 7 LSPPs (LSPP2, LSPP4, LSPP11, LSPP12 (MAP2182c), LSPP12 (MAP2188c), LSPP15, LSPP16) in fecal samples of cattle and buffaloes suspected for Johne’s disease.

The role of TNF receptors as mediators of chronic vasculitis in possible milder forms of the CNS tuberculosis.

Central nervous system (CNS) tuberculosis (TB) is the most severe form of TB due to its high mortality and functional sequelae. There are several differential diagnoses for TB; and, it can also cause secondary conditions, such as vasculitis. 155 biopsies, corresponding to 155 different patients out of 5,386 registered biopsies from 2008-2013, met the criteria of unknown etiology vasculitis and evidence of cerebral vascular disease. These were analyzed to assess the presence of central nervous system TB. The selected cases were assessed with Suzaan Marais (SM) criteria for clinical tuberculosis. After that, Ziehl-Neelsen (ZN) staining and polymerase chain reaction (PCR) were performed to amplify a fragment of the insertion sequence IS6110 of M. tuberculosis. 21 patients met the criteria for definitive tuberculosis by ZN staining and PCR, and 2 met the criteria for possible tuberculosis. Tumor necrosis factor (TNF)-α, TNF-R1, and TNF-R2 were determined by immunohistochemistry in histological sections from formalin-fixed paraffin-embedded (FF-PE) tissues in the 23 selected patients. Granulomatous TB was present in almost half of the cases. TNF-R1 and TNF-R2 were expressed mainly in blood vessels, histiocytes, and macrophages. TNF-R2 expression was higher than the other markers, which suggests an anti-inflammatory response against M. tuberculosis. The histopathological presentation of TB is not always limited to granulomas, abscesses, or meningitis; there are also clinical presentations characterized only with chronic inflammation of nervous and vascular tissue.

The Association Between Vitamin D Levels and Intestinal Parasites

Immune system is known to be affected by vitamin D deficiency. In this study, we investigated whether there was a significant association between vitamin D levels and intestinal parasites. A total of 239 patients treated at the internal medicine outpatient clinics with gastrointestinal complaints were included in this study. Demographic characteristics of the patients were recorded. Examination of stool samples were repeated three times within 10 days. Cellophane band method, native-lugol, sedimentation and modified Kinyoun's acid fast stain methods were used for the detection of parasites in the stool samples. The samples were prepared, stained and examined under a microscope. Vitamin D levels were examined in the blood samples collected from the patients. Patients having vitamin D level under 20 ng/ml were diagnosed with vitamin D deficiency. Binary logistic regression analysis showed that serum vitamin D level was significant informative variable for only Iodamoeba and Cyclospora. No association between vitamin D level and the presence of other intestinal parasites was found. The risk of Iodamoeba positivity was 2.54 times higher in the patients with a serum vitamin D level under 20 ng/ml ($$p<0.01$$). Similarly, the risk of Cyclospora positivity was 2.44 times higher in the patients with a serum vitamin D level under 20 ng/ml compared to those with a serum vitamin D level over 20 ng/ml ($$p<0.01$$). The risk for positivity of Iodamoeba and Cyclospora that are among the intestinal protozoa is high for people with vitamin D deficiency.

Tuberculosis Mimicking-Musculoskeletal Tumor of The Hand: An Uncommon Case of Extrapulmonary Tuberculosis

Background: Musculoskeletal extrapulmonary tuberculosis is accounting for only 10-15% of all cases. Current attention has been focused on hand tuberculosis due to its increasing prevalence. This study describes an unusual case of extrapulmonary tuberculosis that manifested as a soft tissue tumor of the hand.Case Report: A 17-year-old male presented with a painful swollen part of the 3rd finger of the left hand. The mass was irregular and suppurative, with discoloration and foul smell. Laboratory results showed anemia and increased erythrocyte sedimentation rate. The computed tomography scan showed an isodense soft tissue (size 2.19 cm x 1.59 cm x 4.21cm) across the distal to the proximal part of 3rd phalanx bone to the metacarpal region with a decrease of phalanx bone density with no sign of bone destruction or any periosteal reaction. The histopathology study revealed a chronic granulomatous structure surrounded by epitheloid cells, datia Langhans cells, and caseating necrosis. The acid-fast staining confirmed the presence of bacilliform bacteria.Discussion: Extrapulmonary tuberculosis typically results from secondary hematogenous dissemination and reactivation of the main foci. There are no specific tests to diagnose musculoskeletal tuberculosis preoperatively, leading to delayed diagnosis. Histopathological examination and culture are needed to confirm the diagnosis.Conclusions: Slow progression and multiple differential diagnoses frequently result in a delayed diagnosis, which leads to complications. Initial radical excision and anti-tuberculosis multidrug therapy provide excellent functional outcomes and recurrence prevention.