Lowenstein-Jensen Culture Research Articles

The Yield of Sputum Smear Direct Microscopy Using Ziehl-Neelsen Stain in Comparison with Lowenstein Jensen Culture on the Diagnosis of Pulmonary Tuberculosis in Tripoli-Libya

Background: In the developing countries, Tuberculosis is one of leading causes of morbidity and mortality. The detection and identification of Mycobacterium tuberculosis is considered a great challenge for clinical microbiologists. Albeit Ziehl-Neelsen (ZN) smear is being a rapid diagnostic tool for TB, the culture on Lowenstein Jensen (LJ) medium is considered the golden diagnostic tool for TB in the developing countries. Objectives: To evaluate the validity and reliability of ZN smear of acid-fast bacilli compared to the culture on LJ medium. Design and Settings: This study is the first in Libya which was conducted from January 2016 to December 2016 in the TB reference laboratory at the National Center for Disease Control (NCDC), Tripoli, Libya. Patients and methods: The retrospective analytic comparative cross-sectional study included patients visiting TB OPD clinic at NCDC, including transferred patients from other healthcare facilities and hospitals. A total of 2783 specimen of suspected pulmonary TB cases were processed for both direct smear and culture. The direct smears were stained with ZN method (1% Carbolfuchsin, 3% Hydrochloric acid Ethanol and 0.1% methylene blue). Specimens were observed under 100-x oil immersion lens, whilst cultures were inoculated on LJ medium after digestion and decontamination of the clinical specimens. Results: Of total 2783 study subjects, 203 (7.29%) were smear positive while 327 (11.74%) were culture positive. Out of 203 smear positive pulmonary cases, 154 (47.094%) were found to be positive on LJ culture. A total of 2407 pulmonary cases were negative on the smear and LJ culture for Mycobacterium tuberculosis. Conclusion: This preliminary study indicated the low sensitivity of sputum smear direct microscopy for early diagnosis of TB, while culture on LJ remains the golden standard modality.

OMNIgene.SPUTUM suppresses contaminants while maintaining Mycobacterium tuberculosis viability and obviates cold-chain transport.

Tuberculosis (TB) diagnostics are centralised, requiring long-distance transportation of specimens in most resource-limited settings. We evaluated the ability of OMNIgene.SPUTUM (OM-S) to obviate cold-chain transport of TB specimens.A two-arm (same-day and after 5 days sample processing) study was conducted to assess contamination rates and Mycobacterium tuberculosis viability in OM-S-treated samples against the standard decontamination procedure (SDP) in Mozambique, using Lowenstein Jensen (LJ) and mycobacterial growth indicator tube (MGIT) culture and molecular bacterial load assay.270 specimens were processed using OM-S and SDP in same-day and 5-day arms. Contamination was lower in OM-S-treated than SDP-treated cultures: 12% versus 15% and 2% versus 27% in the same-day and 5-day arms, respectively. M. tuberculosis recovery in OM-S-treated LJ cultures was 10% and 56% higher in the same-day and 5-day arms, respectively, than SDP-treated cultures, but lower in MGIT (52% and 28% lower in the same-day and 5-day arms, respectively). M. tuberculosis viable count was 1log estimated CFU·mL−1 lower in 5-day OM-S-treated sputa. OM-S was more effective at liquefying sputum with a shorter sample processing time: 22 min for culture.OM-S is simple to use and has demonstrated a high potency to suppress contaminants, maintenance of viability at ambient temperatures and higher M. tuberculosis recovery, particularly in the solid LJ cultures. Optimisation of OM-S to achieve higher MGIT culture positivity and shorter time to result will increase its application and utility in the clinical management of TB.

Assessment of diagnostic accuracy of GeneXpert Mycobacterium tuberculosis/rifampicin in diagnosis of pulmonary tuberculosis in Kosovo

Background: Despite control program for Mycobacterium tuberculosis (MTB) and anti-tuberculosis (TB) drugs, MTB continues to be a major threat to global health; furthermore, it is the second leading cause of death worldwide. The distribution of TB in Kosovo has the highest rate in Europe. This study was designed to assess the performance of single-sputum GeneXpert MTB/ rifampicin (RIF) assay against the Ziehl-Neelsen smear, solid culture in diagnosing suspected pulmonary cases, and RIF resistance. Methods: A cross-sectional study was conducted to examine various diagnostic methods between 2014 and 2016 in the Department of Microbiology, the National Institute of Public Health, Kosovo, and the Lung Clinic. The present study compares diagnostic methods of MTBC and RIF resistance using Xpert MTB/RIF test in 813 samples from 760 patients suspected of TB by conventional smear microscopy and culture method. Results: Three hundred and ninety-six patients (52.1%) were male and the mean age was 48.6 ± 18.1 years. A total of 261 patients (32.6%) had underlying lung diseases. Of the 801 specimens investigated, 165 (20.5%) were MTBC-positive by culture, while 199 (28.3%) were positive by Xpert assay. Among the examined specimens, Two specimens with false negatives were shown in the Xpert method. Compared with culture, the Xpert assay achieved 82.3% (95% confidence interval [CI]: 65.5%–93.2%) sensitivity and 97.6% (95% CI: 91.5%–99.7%) specificity. GeneXpert could detect 11.7% and 50% additional positive cases as compared to Lowenstein–Jensen culture and smear microscopy, respectively. Three cases with resistance to RIF were detected from clinical isolates. Conclusions: According to the results obtained in this study, GeneXpert MTB/RIF assay can be used as a useful tool for rapid and efficient diagnosis of TB.

The value of nasopharyngeal aspirate, gastric aspirate and bronchoalveolar lavage fluid in the diagnosis of childhood tuberculosis.

Çakır E, Özdemir A, Daşkaya H, Umutoğlu T, Yüksel M. The value of nasopharyngeal aspirate, gastric aspirate and bronchoalveolar lavage fluid in the diagnosis of childhood tuberculosis. Turk J Pediatr 2018; 60: 10-13. Pulmonary tuberculosis (TB) is an important cause of morbidity and mortality especially in developing countries. A definitive microbiologic confirmation of Mycobacterium tuberculosis is important in the diagnosis of childhood TB. We aimed to compare the diagnostic value of nasopharyngeal aspirate (NPA), gastric aspirate (GA) and bronchoalveolar lavage (BAL) specimens in children with highly suspected pulmonary tuberculosis (TB). NPA, GA and BAL samples were obtained from forty patients. The mean age was 9.2±4.7 years. Sixty-eight percent of children had a history of household contact and 82% had tuberculin skin test positivity. Acid-fast bacilli (AFB) stain was positive in 22.5% (N=9) of BAL, 17.5% (N=7) of GA, and 10% (N=4) of NPA samples. Positive Lowenstein-Jensen culture was 27.5% (N=11) in BAL, 22.5% (N=9) in GA, and 12.5% (N=5) in NPA samples. Positive AFB stains and growth in TB cultures from BAL fluid and GA samples were both higher than NPA samples (p < 0.006 and p < 0.004, respectively GA). We conclude that NPA specimen fails to determine Mycobacterium tuberculosis in children with highly suspected pulmonary TB when compared to GA or BAL fluid.

Improved performance of Xpert MTB/RIF assay on sputum sediment samples obtained from presumptive pulmonary tuberculosis cases at Kibong\u2019oto infectious diseases hospital in\xa0Tanzania

BackgroundThe introduction of Xpert MTB/RIF assay (Xpert) has significantly improved diagnosis of Tuberculosis (TB) in resource limited human immunodeficiency virus (HIV) endemic settings. We aimed to modify the Xpert protocol to improve the detection of Mycobacterium tuberculosis (MTB).MethodsThis cross sectional study was conducted among presumptive pulmonary tuberculosis (PTB) patients at Kibong’oto Infectious Diseases Hospital between August and November 2015. Each patient consented to provide 2 samples of raw sputa. One-sputum sample was sedimented using the Petroff’s method and divided into two portions. One portion of sediment was inoculated on Lowenstein-Jensen culture media and observed for any growth for up to 8 weeks. Both, raw sputum and the portions of sediments were tested separately using Xpert with a sample reagent ratio of 1:2. Mean age of patients, prevalence of MTB, Xpert sensitivity, specificity, positive and negative predictive value were calculated. An incremental sensitivity was determined. Pearson chi-square and either an independent T or Mann-Whitney U-test were used to compared categorical and continuous variables respectively. A p- value of ≤0.05 was considered significant.ResultsOf the 270 presumptive PTB cases, 262 were eligible for analysis. Eight (3%) were excluded due to contaminated culture. Patients’ mean age was 42.9 (±SD 15.1) years of which 173 (66%) were female. The overall prevalence of PTB was 112 (43%), of which the Xpert detected 105 (40%) in sediments and 98 (37%) in raw sputa as compared to culture which detected 85 (32%) cases of PTB. Sensitivity, specificity, positive and negative predictive values of Xpert on sputum sediments were 92%, 85%, 74% and 96% respectively. Overall, the incremental sensitivity of Xpert on sediment over raw sputum was 6%. In HIV infected Presumptive PTB, the incremental sensitivity was 12%.ConclusionLowering the sample reagent to sediment dilution ratio increases sensitivity of Xpert on MTB detection among presumptive PTB cases, especially in HIV infected individuals.

The value of microscopic-observation drug susceptibility assay in the diagnosis of tuberculosis and detection of multidrug resistance.

Inexpensive, rapid, and reliable tests for detecting the presence and drug susceptibility of Mycobacterium tuberculosis complex (MTBC) are urgently needed to control the transmission of tuberculosis. In this study, we aimed to assess the accuracy and speed of the microscopic-observation drug susceptibility (MODS) assay in the identification of MTBC and detection of multidrug resistance. Sputum samples from patients suspected to have tuberculosis were simultaneously tested with MODS and conventional culture [Löwenstein-Jensen (LJ) culture, BACTEC MGIT™ 960 (MGIT) system], and drug susceptibility testing (MGIT system) methods. A total of 331 sputum samples were analyzed. Sensitivity and specificity of MODS assay for detection of MTBC strains were 96% and 98.8%, respectively. MODS assay detected multidrug resistant MTBC isolates with 92.3% sensitivity and 96.6% specificity. Median time to culture positivity was similar for MGIT (8days) and MODS culture (8days), but was significantly longer with LJ culture (20days) (p<0.0001 for both comparisons). Median time to availability of the susceptibility results was significantly (p<0.0001) shorter with MODS assay (8days) than MGIT system (20days). In conclusion, MODS is an inexpensive and rapid test with good performance characteristics for direct diagnosis of tuberculosis and detection of multidrug resistance.

An integrated approach to detection of Mycobacterium tuberculosis using BD MGIT 320 system and other diagnostic modalities.

Background: Application of automated liquid culture system like Becton, Dickinson and Company (BD) MGIT 320 can shorten the recovery time of Mycobacteria in clinical diagnostic laboratories as compared to Lowenstein Jensen (LJ) medium. The present study aimed to compare the results of BD MGIT 320 culture system with that of LJ medium and also to evaluate the usefulness of molecular methods as an adjunct. Methods: Specimen from clinically suspected cases of tuberculosis were processed as per standard guidelines for smear microscopy, solid culture in LJ media, liquid culture in BD MGIT 320 system and Line probe assay. Polymerase chain reaction was performed on extracted DNA from MGIT positive specimen to detect IS6110 and MPB64 gene targets. Result: Out of total 83 specimens, a statistically significant higher isolation rate (25.3%, n=21) was shown by MGIT 320 system in comparison to LJ medium (12.04%, n=10). Line Probe Assay was positive in 37.3% (n=31) and direct microscopy was positive in only 13.25% (n=11) specimen. The average duration of detection was found to be more rapid by MGIT320 than LJ culture (18 days vs 24.1 days). Among the 21 MGIT positive specimens 7 were found positive for both IS6110 and MPB64 gene while 7 specimens were found positive only for IS6110 . Conclusion: The automated liquid culture systems like BD MGIT 320 have a higher isolation rate and are more rapid as compared to solid media. But adoption of Line probe assay combined with MGIT culture will definitely yield rapid and efficient detection of Mycobacteria. DOI: 10.21276/AABS.1736

Light-emitting diode fluorescent microscopy and Xpert MTB/RIF\xae assay for diagnosis of pulmonary tuberculosis among patients attending Ambo hospital, west-central Ethiopia

BackgroundThe relatively simple and cheaper light-emitting diode fluorescent microscopy (LED-FM) was recommended by the World Health Organization (WHO) to replace the conventional tuberculosis (TB) microscopy in both high- and low-volume laboratories. More recently the WHO also endorsed one more technique, Xpert MTB/RIF® assay (Xpert), for improved TB diagnosis particularly among human immunodeficiency virus (HIV)-infected cases. However, the relative performance of both of these tools differs from setting to setting in reference to the conventional TB diagnostics. This study thus aimed to evaluate these tools for TB detection in individuals visiting Ambo Hospital, west-central Ethiopia.MethodsCross-sectional early-morning sputum samples were collected from presumptive TB patients between January and August 2015. Socio-demographic data were captured using a structured questionnaire. Clinical information was gathered from patients’ medical records. The sputum samples were diagnosed using LED-FM, Xpert, concentrated Ziehl-Neelsen (cZN) staining and Lowenstein-Jensen (LJ) culture as the gold standard. Drug sensitivity test (DST) was also conducted.ResultsOut of 362 sputum samples collected and processed, 36(9.9%) were positive by LED-FM, 42(11.6%) by cZN and 50(13.8%) by Xpert. But, only 340 samples could be declared culture positive or negative for mycobacteria. Of these 340, eight were non-tubercle mycobacteria (NTM). Out of the remaining 332 samples, 45(13.6%) had culture-confirmed TB with 11(24.4%) being HIV co-infected. LED-FM, Xpert and culture detected 54.5% (6/11), 90.9% (10/11) and 100% (11/11) mycobacteria in HIV-positive individuals and 81.3% (26/32), 73.7% (28/38), 78.8% (26/33) and 73.2% (30/41), in HIV negatives respectively. Two samples were rifampicin resistant by both Xpert and DST. The overall sensitivity, specificity, positive and negative predictive values of LED-FM and Xpert were 77.8, 100, 100 and 96; and 93.3, 98, 97.5 and 98.9% respectively.ConclusionThe data demonstrated the high diagnostic yield of Xpert. LED-FM sensitivity is higher compared to results quoted by recent systematic reviews although it appears to be lower than what was cited in the WHO policy statement (83.6%) during the recommendation of the technology. The high specificity of LED-FM in the study area is encouraging and is expected to boost its reliability and uptake.

Confirmation of positive acid fast bacilli samples by tuberculosis bacilli culture

This study attempts to estimate the efficiency of acid fast bacilli(AFB) stain technique to diagnosis tuberculosis in the different samples. This study was performed in general Al-Diawynia hospital and Department of Microbiology, Medicine College, Al-Qadisiha University between October 2005 to October 2008. A total of 1654 clinical samples from 750 patients analyzed for acid-fast staining. These were 605 sputum, 67 bronchial wash, 42 pleural fluids, 4 urine, 7 ascetic fluids and 3 cerebrospinal fluids. The stained smear-positive were (5.3 %) distributed in different samples .Seventy five patients (10%) were evaluated by Lowenstein-Jensen (LJ) medium culture that it was employed as gold standard for tuberculosis diagnosis, of these 58 were culture-positive specimens. The sensitivity of Ziehl-Neelsen (ZN) stain Methods and LJ culture were (68.9 %) and (95.2 %) respectively. The specificity of ZN stain Methods and LJ culture were (88.2 %) and (45.4 %) respectively. The diagnosis accuracy of ZN stain Methods and LJ culture were (73.3 %) and (73.3 %) respectively. The positive predictive value of ZN stain Methods and LJ culture were (95.2 %) and (68.9 %), while negative predictive value were (45.4 %) and (88.2 %) respectively.

GeneXpert technology. A breakthrough for the diagnosis of tuberculous pericarditis and pleuritis in less than 2 hours.

Objectives:To evaluate the diagnostic validity of GeneXpert for the detection of Mycobacterium tuberculosis (MTB) in pericardial and pleural effusions samples.Methods:A cross sectional study was conducted at the Mycobacteriology Laboratory, Allama Iqbal Medical College, Lahore, Pakistan. A total of 286 (158 pleural and 128 pericardial fluids) samples were received from tuberculosis (TB) suspects between January 2014 and August 2016. Every sample was processed for Ziehl-Neelsen (Zn) smear, Lowenstein Jensen (LJ) culture, GeneXpert MTB/RIF assay according to standard protocols. Validity of GeneXpert assay for the detection of MTB was evaluated using LJ culture as gold standard.Results:Out of 286 effusions samples, MTB was isolated by LJ culture in 51 (17.8%) samples followed by GeneXpert in 43 (15%), and acid- fast bacilli (AFB) was detected by Zn smear microscopy in 11 (3.8%) samples. GeneXpert showed high sensitivity (84.3%), specificity (100%), with positive predictive value (100%), and negative predictive value (96.7%), while Zn smear showed sensitivity 18.3%, specificity 99.1%, positive predictive value 81.8%, and negative predictive value 85.4%. A strikingly high sensitivity of 72.2% was observed for pericardial fluid by GeneXpert.Conclusion:GeneXpert assay is an innovative tool, for prompt detection of MTB and drug resistance. It is definitely an attractive point of care test, with high sensitivity and specificity along with turn around time of 2 hours, which facilitates timely diagnosis and appropriate management of TB pleuritis and pericarditis.

Rapid and Specific Diagnosis of Extrapulmonary Tuberculosis by Immunostaining of Tissues and Aspirates With Anti-MPT64.

Extrapulmonary tuberculosis (EPTB) constitutes about 15% to 20% of all cases of tuberculosis (TB). The confirmation of EPTB has always been a challenge to laboratory personnel. We aim to evaluate the diagnostic potential of immunostaining with anti-MPT64 in various EPTB specimens. We studied a total of 51 TB cases and 38 non-TB control specimens comprising of fine-needle aspirates and formalin-fixed biopsies. These were investigated using a combination of the Ziehl-Neelsen method, the Lowenstein-Jensen culture, immunostaining with anti-MPT64 and anti-BCG, and nested-polymerase chain reaction (PCR) for IS6110. Results of all the tests were compared using nested-PCR as the gold standard. Diagnostic validation of immunostaining for anti-MPT64 was performed using nested-PCR as the gold standard. The overall sensitivity, specificity, and positive and negative predictive values for immunostaining with anti-MPT64 were 100%, 97%, 97%, and 100%, respectively. Immunostaining using anti-MPT64 is a rapid and sensitive method for establishing an early and specific diagnosis of Mycobacterium tuberculosis infection. The technique is simple to be incorporated into routine pathology laboratories.

Use of DNA microarray chips for the rapid detection of Mycobacterium tuberculosis resistance to rifampicin and isoniazid.

The objective of the present study was to evaluate the potential development of DNA microarray chips to detect rifampicin (RFP) and isoniazid (INH) resistance in Mycobacterium tuberculosis (MTB), using samples from clinical tuberculosis (TB) patients in Soochow City, China. The sputum samples of 42 patients with TB in the Affiliated Hospital of Infectious Diseases of Soochow University (Soochow, China) were collected. The conventional Lowenstein-Jensen culture medium (Gold Standard) was used to assess drug sensitivity using the absolute concentration method. GeeDom MTB drug detection kits were also used to create a DNA microarray chip and examine the RFP-resistance associated gene mutation points rpoB-RRDR 511, 513, 516, 526, 531 and 533, and the INH-resistance associated gene mutation points katG315 and inhA-15 of the sputum samples. Compared with the results from the absolute concentration method, the susceptibility and specificity of RFP sensitivity detected by the DNA microarray chip were 92.8 and 93.8%, respectively. The susceptibility and specificity of INH sensitivity detected were 66.7 and 81%, respectively. The rpoB-RRDR 526, 531 mutations were the primary causes of MTB RFP resistance and the katG315 mutation was the primary cause of INH resistance. The detection of rpoB and katG gene mutation points by a DNA microarray chip may be used as a rapid, accurate and bulk clinical detection method for RFP and INH resistance in MTB. This is very valuable for the control of TB epidemics.

Screening for genital tuberculosis in a limited resource country: case report

BackgroundScreening for benign or malignant process of pelvis in young females is a challenge for a physician in a limited resource country. Tuberculosis should be always considered in the differential diagnosis of a pelvic mass in countries with high prevalence of tuberculosis. Negative results of analysis of peritoneal fluid for acid-fast staining, late cultures, and unavailability of new diagnostics methods such as polymerase chain reaction and adenosine deaminase of the aspirated fluid from peritoneal cavity can often result in invasive diagnostic procedures such as laparotomy.Case presentationWe report a case of a 24 year old Albanian unemployed female living in urban place in Kosovo who presented with abdominal pain, loss of appetite, fever, headache, a weight loss, nonproductive cough and menstrual irregularity for three weeks. In this example case, the patient with cystic mass in tubo-ovarial complex and elevated serum cancer antigen 125 levels was diagnosed for genital tuberculosis after performing laparotomy. Caseose mass found in left tubo-ovarial complex and histopathological examination of biopsied tissue were the fastest diagnostic tools for confirming pelvis TB. The Lowenstein-Jensen cultures were positive after six weeks and her family history was positive for tuberculosis.ConclusionYoung females with abdominopelvic mass, ascites, a positive family history for tuberculosis and high serum cancer antigen 125, should always raise suspicion of tuberculosis especially in a limited resource country. A laparoscopy combined with peritoneal biopsy should be performed to confirm the diagnosis as this could lead to a prevention of unnecessary laparotomies.

Investigation of OMNIgene·SPUTUM performance in delayed tuberculosis testing by smear, culture, and Xpert MTB/RIF assays in Uganda

OMNIgene·SPUTUM (OM-S) is a sample transport reagent designed to work with all tuberculosis diagnostics while eliminating the need for cold chain. OM-S-treated sputum samples were assayed in several tests after multiday holds. Raw sputa from 100 patients underwent direct smear microscopy, were manually split and assigned to the OM-S group [OM-S added at collection (no other processing required) and tested after 0- to 5-day holds at room temperature] or standard-of-care (SOC) group (NaOH/N-acetyl l-cysteine decontamination, all tested on day of collection). Concentrated smear microscopy, Lowenstein Jensen (LJ) culture, and mycobacteria growth indicator tube (MGIT) culture were performed. For patients with negative direct smear, a second sample was split, with SOC (raw sputum) and OM-S portions (sediment) tested in the Xpert MTB/RIF (Xpert) assay. OM-S group and SOC group results were strongly concordant on all four tests [range, 89% (MGIT)–97% (Xpert)]. OM-S MGIT, LJ, and Xpert tests were in statistical agreement with SOC MGIT as reference. OM-S specimens had lower culture contamination rates (3% vs. 10% LJ; 2% vs. 5% MGIT) but required, on average, 5.6 additional days to become MGIT-positive. The findings suggest that samples held/transported in OM-S are compatible with smear microscopy, LJ or MGIT culture, and Xpert, and perform comparably to fresh sputum samples. Larger feasibility studies are warranted.

Evaluation of Xpert MTB/RIF assay performance in the diagnosis of abdominal tuberculosis.

Background/AimsThe use of genetic probes for the diagnosis of pulmonary tuberculosis (TB) has been well described. However, the role of these assays in the diagnosis of intestinal tuberculosis is unclear. We therefore assessed the diagnostic utility of the Xpert Mycobacterium tuberculosis/rifampicin (MTB/RIF) assay, and estimated the prevalence of multidrug-resistant (MDR) TB in the Indian population.MethodsOf 99 patients recruited, 37 had intestinal TB; two control groups comprised 43 with Crohn's disease (CD) and 19 with irritable bowel syndrome. Colonoscopy was performed before starting any therapy; mucosal biopsies were subjected to histopathology, acid-fast bacilli staining, Lowenstein-Jensen culture, and nucleic acid amplification testing using the Xpert MTB/RIF assay. Patients were followed up for 6 months to confirm the diagnosis and response to therapy. A composite reference standard was used for diagnosis of TB and assessment of the diagnostic utility of the Xpert MTB/RIF assay.ResultsOf 37 intestinal TB patients, the Xpert MTB/RIF assay was positive in three of 37 (8.1%), but none had MDR-TB. The sensitivity, specificity, positive predictive value, and negative predictive value of the Xpert MTB/RIF assay was 8.1%, 100%, 100%, and, 64.2%, respectively.ConclusionsThe Xpert MTB/RIF assay has low sensitivity but high specificity for intestinal TB, and may be helpful in endemic tuberculosis areas, when clinicians are faced with difficulty differentiating TB and CD. Based on the Xpert MTB/RIF assay, the prevalence of intestinal MDR-TB is low in the Indian population.

An alert of Mycobacterium tuberculosis infection of rhesus macaques in a wild zoo in China.

Mycobacterium tuberculosis, the pathogen that causes tuberculosis (TB), is becoming increasingly recognized as an important cause of fatal chronic illnesses in China. In this study, we report an infectious disease among 84 rhesus macaques at a Chinese zoo. Their clinical signs and symptoms were very similar with the manifestations of TB in humans. To determine the potential pathogens of this outbreak, many methods were used. First, tuberculin skin tests showed that none of the monkeys displayed significant skin reactions. Subsequently, the sera were tested for specific antibody IgG; 29 (34.5%) and 39 (46.4%) blood samples tested positive by TB-IgG and TB-DOT, respectively. Radiographic examination showed characteristic imageology changes in 14 (16.7%) monkeys. One individual determined as positive by the above three methods was euthanized, and histopathological analysis demonstrated typical granulomas and caseous necrosis in the lung, liver, spleen, and intestine. Furthermore, the pathogenic mycobacteria were isolated from lung lobe, cultured on acidic Lowenstein-Jensen culture medium, and identified as M. tuberculosis by real-time PCR and DNA sequencing. Nevertheless, the origin of the infection remained unknown. These findings emphasize the need to strengthen the management and training of staff, especially those working at animal shelters.

Thin-layer agar (TL7H11) for rapid isolation of &lt;i&gt;Mycobacterium tuberculosis&lt;/i&gt; in sputum specimens

Background: Tuberculosis (TB) remains one of the major causes of death from a single infectious agent worldwide. The early detection of new cases of pulmonary tuberculosis is an important goal in tuberculosis control program.Objective: 1n this study, thin layer agar (TLA) culture was compared with Lowenstein-Jensen (LJ) culture for rapid detection of pulmonary tuberculosis. Methods: It was a cross sectional study conducted in National Tuberculosis Reference Labora­tory (NTRL) of National Institute of Disease of Chest and Hospital (NIDCH), Dhaka, from July 2010 to June 2011. A total of 100 sputum smear positive for acid fast bacilli (AFB) by Z-N staining, pulmonary tuberculosis patients were included in this study. Samples were processed by modified Petroff method and then cultured on thin layer 7H11(TL7H11) plates and L-J tubes. TL7H11 plates were observed microscopically for rnicrocolony growth once a week for 6 weeks, and L-J tubes were observed once a week for 8 weeks. Results: The recovery rates of mycobacteria on only TLA, only LJ and on both media were 90%, 97% and 88% respectively. Overall positivity was 99% in both L-J and TLA media. Mean time for detection of mycobacteria on TLA was 9.04±1.66 days compared to 21.78±6.19 days on L-J media. The rate of contamination was higher (6%) in L-J media than in TLA media (4%). Conclusion: The TL7H11 media can be used as an alternative to the Lowenstein-Jensen medium for early isolation of mycobacteria in resource constrained settings.

Primary and secondary anti-tuberculosis drug resistance in Hitossa District of Arsi Zone, Oromia Regional State, Central Ethiopia.

BackgroundMultidrug-resistant tuberculosis (MDR-TB) drugs which is resistant to the major first-line anti-TB drugs, Isoniazid and Rifampicin, has become a major global challenge in tuberculosis (TB) control programme. However, its burden at community level is not well known. Thus, the aim of study was to assess the prevalence of primary and secondary resistance to any first line anti-TB drugs and MDR TB in Hitossa District of Oromia Regional State, Central Ethiopia.MethodsPopulation based cross- sectional study was conducted on individuals aged ≥15 years. Those with symptoms suggestive of TB were interviewed and two sputum specimens were collected from each and examined using Lowenstein-Jensen (LJ) culture medium. Further, the isolates were confirmed by the Ziehl-Neelsen microscopic examination method. Drug susceptibility test (DST) was also conducted on LJ medium using a simplified indirect proportion method. The resistance strains were then determined by percentage of colonies that grew on the critical concentration of Isoniazid, Streptomycin, Rifampicin and Ethambutol.ResultsThe overall resistance of all forms of TB to any first-line anti-TB drug was 21.7 %. Of the total new and previously treated culture positive TB cases, 15.3 and 48.8 % respectively were found to be a resistant to any of the first-line anti-TB drugs. Further, of all forms of TB, the overall resistance of MDR-TB was 4.7 %. However, of the total new TB cases, 2.4 % had primary while 14.3 % had secondary MDR-TB. Resistance to any of the first-line anti-TB drugs (adjusted odd ratio (AOR), 8.1; 95 % CI: 2.26–29.30) and MDR-TB (AOR), 7.1; 95 % CI: 2.6–43.8) was found to be linked with previous history of anti-TB treatment.ConclusionsThe study has identified a high rate of primary and secondary resistance to any of the first-line anti-TB drugs and MDR-TB in the study area. The resistance may have resulted from sub-optimal performance of directly observed treatment short-course (DOTS) programme in the detecting infectious TB cases and cure rates in the study area. Anti-TB drug resistance is linked with previous TB treatment. There is a need to strengthen DOTS and DOTS-Plus programmes and expand MDR-TB diagnostic facilities in order to timely diagnose MDR-TB cases and provide appropriate treatment to prevent the spread of MDR-TB in Ethiopia.Electronic supplementary materialThe online version of this article (doi:10.1186/s12889-016-3210-y) contains supplementary material, which is available to authorized users.

Loop-mediated isothermal amplification assay for detection of Mycobacterium tuberculosis complex in infertile women

Background: Female genital tuberculosis (FGTB) has a profound impact on the reproductive health of patients including infertility. Conventional diagnostic techniques have low sensitivity and specificity as well as long turnaround time. There is a need of developing newer, rapid and practically adaptable technique, especially in low-income countries. Objective: To standardize and evaluate loop-mediated isothermal amplification (LAMP) technique for diagnosis of FGTB. Methods: A total of 300 endometrial biopsy samples from infertile females were subjected to Ziehl–Neelsen (ZN) staining, Lowenstein–Jensen culture, automated culture (BACTEC mycobacterial growth indicator tube), histopathological examination (HPE), nucleic acid amplification by polymerase chain reaction (PCR) and LAMP technique. Composite gold standard (either smear/culture/HPE/PCR positive) was considered for calculation of outcome parameters. Results: The observed sensitivities of ZN smear, culture, HPE, PCR and LAMP were 2.94%, 10.29%, 8.82%, 95.59% and 66.18%, respectively. Overall concordance between PCR and LAMP was 63%, which shows a good agreement. Conclusion: This study is the first to evaluate LAMP in the diagnosis of FGTB and found it to be a rapid and convenient technique, especially in low resource endemic settings.

An Unusual Cause of Dysphagia with Bronchoesophageal Fistula

A 57-year-old man presented with a 4-week history of dysphagia and cough. Physical examination revealed mucocutaneous pallor, and laboratory tests showed a slight anemia (hemoglobin level 12,5 g/dL). An upper gastrointestinal endoscopy reveled an obstructive neoplasm in the midesophagus (Fig. 1); biopsy specimens showed fibrinonecrotic and granulation tissue. Chest computed tomography showed a large peribronchial mass with bronchoesophageal fistula (BEF) (Fig. 2), and bronchoscopy reveled bronchial infiltration and marked caliber reduction of the left main bronchus. The results of examinations of samples from repeat biopsies of esophageal and bronchial lesions were negative for dysplasia or neoplasia. The results of polymerase chain reaction testing for Mycobacterium tuberculosis were positive, so we began administration of tuberculostatics and systemic corticosteroids, assuming lymph node tuberculosis with mediastinal involvement. However, there was a worsening of dyspnea and weight loss, and the results of the Xpert MTB/RIF Assay (Cepheid, Sunnyvale, CA) and Lowenstein–Jensen culture were both negative, leading to the assumption of a false-positive result of the first test. Endoscopic reassessment showed increase esophageal lesion and extensive tracheobronchial destruction, forming a large necrotic cavity in the mediastinum (Fig. 3). Histopathological analysis revealed large lymphocytes with a diffuse growth pattern with negative immunostaining for cytokeratin AE1, cytokeratin AE3, and for cluster of