In Vitro Effect of 5-Nitroimidazole Drugs against Trichomonas vaginalis Clinical Isolates.

Infections with the sexually transmitted parasite Trichomonas vaginalis are normally treated with metronidazole, but cure rates are suboptimal and recurrence rates following treatment are high. Therefore, our objective was to assess the in vitro antitrichomonas activities of three other 5-nitroimidazole drugs and compare them with metronidazole. T. vaginalis isolates (n = 94) isolated from South African women presenting with vaginal discharge syndrome at two sexually transmitted disease clinics in KwaZulu-Natal were grown from frozen stock. Twofold serial dilutions (16 to 0.25 mg/L) of metronidazole, tinidazole, ornidazole, and secnidazole were prepared in Diamond's broth. The MICs were read after 48 h of anaerobic incubation at 37°C. An MIC of <2 mg/L was defined as susceptible, an MIC of 2 mg/L was defined as intermediate, and an MIC of >2 mg/L was defined as resistant. Sixty-one percent (57/94) of the T. vaginalis isolates were susceptible to metronidazole, 80% (75/94) were susceptible to tinidazole, 75% (71/94) were susceptible to secnidazole, and 89% (84/94) were susceptible to ornidazole. Resistance levels were 11%, 2%, and 1% for metronidazole, tinidazole, and secnidazole, respectively, while no resistance was observed for ornidazole. Intermediate scores were 28% for metronidazole, 18% for tinidazole, 24% for secnidazole, and 11% for ornidazole. Isolates from a proportion of women with bacterial vaginosis (BV) had higher MICs, and no isolates from women coinfected with another sexually transmitted infectious organism were resistant to any of the antimicrobials tested. This study showed that among T. vaginalis isolates in KwaZulu-Natal, there is no in vitro resistance to ornidazole. Of the 5-nitroimidazoles, metronidazole showed the highest level of resistance. The very low levels of resistance for the other three antimicrobials indicate that all three are viable options as a replacement for metronidazole if these in vitro findings are found to correlate with clinical outcomes. IMPORTANCE Trichomonas vaginalis is the most common nonviral sexually transmitted infection associated with reproductive sequelae and HIV acquisition risk worldwide. Despite its role in reproductive health, a high prevalence in South Africa, and the reported metronidazole resistance worldwide, no alternative regimens have been tested against T. vaginalis in our setting. This study compared the susceptibility patterns of three other 5-nitroiminazoles (secnidazole, tinidazole, and ornidazole), which are active against T. vaginalis with metronidazole in vitro. Metronidazole, the drug of choice for the treatment of trichomoniasis, showed the highest level of resistance, while the three regimens showed very low levels of resistance. These data indicate that all three are viable options as a replacement for metronidazole if these in vitro findings are found to correlate with clinical outcomes.

Affinity of Mycobacterium tuberculosis strains for M059K microglial cells after migration through A549 alveolar epithelium.

Tuberculosis (TB) remains a major threat worldwide while central nervous system TB (CNS-TB) is one of the most severe forms of extrapulmonary TB. CNS-TB develops as a secondary infection during the hematogenous spread of Mycobacterium tuberculosis (M. tuberculosis) from the lungs to the CNS. Factors influencing the dissemination of the bacilli to the CNS have not been studied extensively. This study evaluated the transmigration ability through the alveolar epithelium and adhesion and invasion capacity of glial cells of M. tuberculosis strains of varying drug susceptibility and genotype profiles using an in vitro co-culture model. A549 alveolar epithelial cells and M059K glial cells were co-cultured in a Transwell plate with A549 cells cultured in the upper chamber and M059K glial cells in the lower chamber. A549 epithelial cells were infected with F15/LAM4/KZN (susceptible, MDR, XDR), Beijing (susceptible, XDR), F11 (susceptible), F28 (MDR), and H37Rv strains of M. tuberculosis. The transmigration of an A549 monolayer and subsequent adhesion and invasion rates of M059K cells were established. The susceptible and XDR variants of the F15/LAM4/KZN strain transmigrate the alveolar epithelial cell monolayer more efficiently than the MDR variant. The Beijing-XDR variant showed a high transmigration rate, while the susceptible variant showed no transmigration ability. Similar to the MDR F15/LAM4/KZN, the F28 and F11 strains showed a low dissemination ability. The bacteria were still capable to adhere to M059K glial cells after passage through the A549 cells. We conclude that M. tuberculosis isolates that passed through a monolayer of A549 alveolar epithelium by transcellular migration can still adhere to M059K glial cells. There is no genetic link between resistance and transmigration.

Stability of a monovalent rotavirus vaccine after exposure to different temperatures observed in KwaZulu-Natal, South Africa.

BACKGROUND: Rotavirus infection and its associated hospitalization of children less than 5 years old in middle- and low-income countries remains a public health challenge. We hypothesized that the Rotarix®potency is affected by non-optimal temperatures which translates into reduced vaccine effectiveness in these settings. OBJECTIVE: To assess the effect of non-optimal temperatures on the potency of the Rotarix® vaccine in South Africa. METHODS: Rotarix® vaccine was exposed to temperatures reflecting breaches in the cold chain. Vero cells (ATCC CCL-81) grown in a 24-well tissue culture plates were infected with Rotarix® vaccine viruses after exposure to non-optimal temperatures and the potency of the vaccine was determined using the plaque assay. RESULTS: Exposure of the Rotarix® vaccine to seasonal temperatures in KwaZulu-Natal for 6 hours and to extreme temperatures of 40oC for 72 hours as well as to -20°C and -80°C for 12 hours did not affect the potency of the vaccine beyond its expected standard of >7 x 105 PFU/ml. CONCLUSION: This study revealed that the Rotarix® vaccine remains potent even after exposure to non-optimal temperatures. However, this study only explored the effect of a constant 'adverse' temperature on vaccine potency and not the effect of temperature fluctuations.

An evaluation of infection control in private and public sector intensive care units in South Africa.

BACKGROUND: Appropriate infection control policies and practices are key to reducing the risk of healthcare-associated infections in patients in intensive care units (ICUs). OBJECTIVE: To evaluate infection control in ICUs using the Infection Control Assessment Tool (ICAT). METHODS: Six public and five private adult ICUs were included. Seven modules from the ICAT were administered including ICU, hand hygiene, and isolation and standard precautions. Modules were scored on a quantitative scale as per the tool guidelines and trained independent nurses observed practices. RESULTS: All ICUs reported to have a 1:1 nurse-to-patient ratio. One public ICU did not have the required 1:2 hand wash basin-to-bed ratio. We observed 100% adherence to handwashing or alcohol rub at each of the five moments of hand hygiene; however, the correct amount of alcohol rub was used in only 2% (n = 2) of the 117 observations. The median score for isolation and standard precautions was 79%. DISCUSSION: There was good infection control practice in ICUs. However, ICUs did not have isolation policies for all the infections listed in the ICAT and did not screen visitors to the ICU. We identified shortcomings in the ICAT and a more suitable tool is required for our healthcare setting.

Challenges with Surveillance of Healthcare-Associated Infections in Intensive Care Units in South Africa.

BACKGROUND: The incidence of healthcare-associated infections (HAIs) in the public health sector in South Africa is not known due to the lack of a surveillance system. We report on the challenges experienced in the implementation of a surveillance system for HAIs in intensive care units (ICUs). METHODS: A passive, paper-based surveillance system was piloted in eight ICUs to measure the incidence of ventilator-associated pneumonia, catheter-associated urinary tract infection, and central line-associated bloodstream infection. Extensive consultation with the ICU clinical and nursing managers informed the development of the surveillance system. The Plan-Do-Study-Act method was utilized to guide the implementation of the surveillance. RESULTS: The intended outputs of the surveillance system were not fully realized due to incomplete data. The organizational culture did not promote the collection of surveillance data. Nurses felt that the surveillance form added to their workload, and the infection control practitioners were unable to adequately supervise the process due to competing work demands. CONCLUSIONS: A manual system that adds to the administrative workload of nurses is not an effective method of measuring the burden of HAIs. Change management is required to promote an organizational culture that supports accurate data collection for HAIs.

Mechanisms of first-line antimicrobial resistance in multi-drug and extensively drug resistant strains of Mycobacterium tuberculosis in KwaZulu-Natal, South Africa.

BACKGROUND: In South Africa, drug resistant tuberculosis is a major public health crisis in the face of the colossal HIV pandemic. METHODS: In an attempt to understand the distribution of drug resistance in our setting, we analysed the rpoB, katG, inhA, pncA and embB genes associated with resistance to key drugs used in the treatment of tuberculosis in clinical isolates of Mycobacterium tuberculosis in the KwaZulu-Natal province. RESULTS: Classical mutations were detected in the katG, inhA and embB genes associated with resistance to isoniazid and ethambutol. Diverse mutations were recorded in the multidrug resistant (MDR) and extensively drug resistant (XDR) isolates for the rpoB and pncA gene associated with resistance to rifampicin and pyrazinamide. CONCLUSIONS: M.tuberculosis strains circulating in our setting display a combination of previously observed mutations, each mediating resistance to a different drug. The MDR and XDR TB isolates analysed in this study displayed classical mutations linked to INH and EMB resistance, whilst diverse mutations were linked to RIF and PZA resistance. The similarity of the XDR strains confirms reports of the clonality of the XDR epidemic. The successful dissemination of the drug resistant strains in the province underscores the need for rapid diagnostics to effectively diagnose drug resistance and guide treatment.

Strains of Mycobacterium tuberculosis differ in affinity for human osteoblasts and alveolar cells in vitro.

Although the lung is the primary site of infection of tuberculosis, Mycobacterium tuberculosis is capable of causing infection at other sites. In 5-10 % such extra-pulmonary tuberculosis is located in bone tissue of the spine. It is unknown whether host or microbial factors are responsible for the site where extra-pulmonary tuberculosis manifests itself. One MDR isolate belonging to strain F28, one susceptible F11 and one isolate each of susceptible, MDR and XDR F15/LAM4/KZN were cultured in Middlebrook 7H9 media. Human osteoblasts (SaOS-2) and human alveolar epithelial cells (A549) were exposed to these different isolates of M. tuberculosis and invasion capacity and intra-cellular multiplication rates were established. Mouse macrophage (MHS) cells exposed to M. tuberculosis H37Rv served as control. The invasion capacity of F15/LAM4/KZN representatives increased with the level of resistance. The F28 MDR strain showed similar invasion capacity as the XDR F15/LAM4/KZN for pulmonary epthelial cells, whilst the fully susceptible F11 strain displayed a propensity for osteoblasts. The differences observed may in part explain why certain strains are able to cause infection at specific extra-pulmonary sites. We postulated that the development of extra-pulmonary tuberculosis depends on the ability of the microbe to pass effectively through the alveolar epithelial lining and its affinity for cells other than those in pulmonary tissue.

Moxifloxacin resistance in the F15/LAM4/KZN extensively drug-resistant strain of Mycobacterium tuberculosis.

OBJECTIVES: Moxifloxacin (MXF) has been advocated for the treatment of extensively drug-resistant (XDR) tuberculosis despite resistance to older-generation fluoroquinolones. We investigated the relationship between the minimum inhibitory concentration (MIC) of MXF and mutations in the gyrA and gyrB genes in Mycobacterium tuberculosis (MTB) isolates from KwaZulu-Natal (KZN) Province of South Africa. MATERIALS AND METHODS: MICs of 56 MTB isolates were compared to the mutations in the quinolone resistance-determining region known to confer fluoroquinolone resistance. Isolates were genotyped by IS6110 restriction fragment length polymorphism analysis. RESULTS: The circulating F15/LAM4/KZN XDR strain circulating in KZN Province harbored the A90V mutation and displayed high-level resistance with MICs of 8 mg/L for ciprofloxacin and ofloxacin and ≥1 mg/L for MXF. CONCLUSION: The inclusion of MXF in XDR-TB treatment regimens requires careful consideration in our setting, where clinical outcome data in MXF-containing regimens are unavailable.

Reduced virulence of an extensively drug-resistant outbreak strain of Mycobacterium tuberculosis in a murine model.

Bacterial drug resistance is often associated with a fitness cost. Large outbreaks of multidrug-resistant (MDR) and extensively drug-resistant (XDR) TB have been described that predominately affect persons with HIV infection. We obtained four closely-related Mycobacterium tuberculosis strains (genotype F15/LAM4/KZN) from an outbreak in KwaZulu-Natal (KZN), South Africa, including drug-sensitive, MDR, and XDR clinical isolates. We compared the virulence of these strains in a murine model of aerosol M. tuberculosis infection for four phenotypes: (1) competitive in vivo growth in lung and spleen, (2) non-competitive in vivo growth in lung and spleen, (3) murine survival time, and (4) lung pathology. When mixtures of sensitive, MDR, and XDR KZN strains were aerosolized (competitive model), lung CFUs were similar at 60 days after infection, and spleen CFUs were ordered as follows: sensitive > MDR > XDR. When individual strains were aerosolized (non-competitive model), modest differences in lung and spleen CFUs were observed with the same ordering. C57BL/6, C3H/FeJ, and SCID mice all survived longer after infection with MDR as compared to sensitive strains. SCID mice infected with an XDR strain survived longer than those infected with MDR or sensitive strains. Lung pathology was reduced after XDR TB infection compared to sensitive or MDR TB infection. In summary, increasing degrees of drug resistance were associated with decreasing murine virulence in this collection of KZN strains as measured by all four virulence phenotypes. The predominance of HIV-infected patients in MDR and XDR TB outbreaks may be explained by decreased virulence of these strains in humans.

Minimal diversity of drug-resistant Mycobacterium tuberculosis strains, South Africa.

Multidrug- (MDR) and extensively drug-resistant tuberculosis (XDR TB) are commonly associated with Beijing strains. However, in KwaZulu-Natal, South Africa, which has among the highest incidence and mortality for MDR and XDR TB, data suggest that non-Beijing strains are driving the epidemic. We conducted a retrospective study to characterize the strain prevalence among drug-susceptible, MDR, and XDR TB cases and determine associations between strain type and survival. Among 297 isolates from 2005-2006, 49 spoligotype patterns were found. Predominant strains were Beijing (ST1) among drug-susceptible isolates (27%), S/Quebec (ST34) in MDR TB (34%) and LAM4/KZN (ST60) in XDR TB (89%). More than 90% of patients were HIV co-infected. MDR TB and XDR TB were independently associated with mortality, but TB strain type was not. We conclude that, although Beijing strain was common among drug-susceptible TB, other strains predominated among MDR TB and XDR TB cases. Drug-resistance was a stronger predictor of survival than strain type.

Nosocomial transmission of extensively drug-resistant tuberculosis in a rural hospital in South Africa.

BACKGROUND: Extensively drug-resistant tuberculosis (XDR-tuberculosis) is a global public health threat, but few data exist elucidating factors driving this epidemic. The initial XDR-tuberculosis report from South Africa suggested transmission is an important factor, but detailed epidemiologic and molecular analyses were not available for further characterization. METHODS: We performed a retrospective, observational study among XDR-tuberculosis patients to identify hospital-associated epidemiologic links. We used spoligotyping, IS6110-based restriction fragment-length polymorphism analysis, and sequencing of resistance-determining regions to identify clusters. Social network analysis was used to construct transmission networks among genotypically clustered patients. RESULTS: Among 148 XDR-tuberculosis patients, 98% were infected with human immunodeficiency virus (HIV), and 59% had smear-positive tuberculosis. Nearly all (93%) were hospitalized while infectious with XDR-tuberculosis (median duration, 15 days; interquartile range: 10-25 days). Genotyping identified a predominant cluster comprising 96% of isolates. Epidemiologic links were identified for 82% of patients; social network analysis demonstrated multiple generations of transmission across a highly interconnected network. CONCLUSIONS: The XDR-tuberculosis epidemic in Tugela Ferry, South Africa, has been highly clonal. However, the epidemic is not the result of a point-source outbreak; rather, a high degree of interconnectedness allowed multiple generations of nosocomial transmission. Similar to the outbreaks of multidrug-resistant tuberculosis in the 1990s, poor infection control, delayed diagnosis, and a high HIV prevalence facilitated transmission. Important lessons from those outbreaks must be applied to stem further expansion of this epidemic.

Mycobacterium tuberculosis isolates grown under oxygen deprivation invade pulmonary epithelial cells.

Mycobacterium tuberculosis has the ability to adapt to and survive under different environmental conditions, including oxygen deprivation. To better understand the pathogenesis of M. tuberculosis, we studied the invasion of human alveolar (A549) and human bronchial (BBM) epithelial cell lines by M. tuberculosis isolates cultured under oxygen deprivation. We used isolates belonging to the Beijing and F15/LAM4/KZN families, isolates with unique DNA fingerprints and the laboratory strains H37Rv and H37Ra. We determined that: (1) M. tuberculosis bacilli grown under oxygen deprivation invade epithelial cells, (2) the invasion capacity of all 17 isolates differed, and (3) oxygen deprivation influenced the invasion capacity of these isolates. All isolates invaded the A549 more effectively than the BBM cells. Three of the F15/LAM4/KZN isolates, two of which had extensively drug resistance (XDR) profiles, were at least twice as invasive (≥33%) as the most invasive Beijing isolate (15%) (P < 0.05). We conclude that for a more comprehensive understanding of the pathogenesis of M. tuberculosis, studies should include isolates that have been cultured under oxygen deprivation.

Symptomatic vaginal discharge is a poor predictor of sexually transmitted infections and genital tract inflammation in high-risk women in South Africa.

BACKGROUND: Diagnosis and treatment of sexually transmitted infections (STIs) is a public health priority, particularly in regions where the incidence of human immunodeficiency virus (HIV) infection is high. In most developing countries, STIs are managed syndromically. We assessed the adequacy of syndromic diagnosis of STIs, compared with laboratory diagnosis of STIs, and evaluated the association between STI diagnosis and the risk of HIV acquisition in a cohort of high-risk women. METHODS: HIV-uninfected high-risk women (n = 242) were followed for 24 months. Symptoms of STIs were recorded, and laboratory diagnosis of common STI pathogens was conducted every 6 months. Forty-two cytokines were measured by Luminex in cervicovaginal lavage specimens at enrollment. Human immunodeficiency virus type 1 (HIV-1) infection was evaluated monthly. RESULTS: Only 12.3% of women (25 of 204) who had a laboratory-diagnosed, discharge-causing STI had clinically evident discharge. Vaginal discharge was thus a poor predictor of laboratory-diagnosed STIs (sensitivity, 12.3%; specificity, 93.8%). Cervicovaginal cytokine concentrations did not differ between women with asymptomatic STIs and those with symptomatic STIs and were elevated in women with asymptomatic STIs, compared with women with no STIs or bacterial vaginosis. Although laboratory-diagnosed STIs were associated with increased risk of HIV infection (hazard ratio, 3.3 [95% confidence interval, 1.5-7.2)], clinical symptoms were not. CONCLUSIONS: Syndromic STI diagnosis dependent on vaginal discharge was poorly predictive of laboratory-diagnosed STI. Laboratory-diagnosed STIs were associated with increased susceptibility to HIV acquisition, while vaginal discharge was not.

φ(2)GFP10, a high-intensity fluorophage, enables detection and rapid drug susceptibility testing of Mycobacterium tuberculosis directly from sputum samples.

The difficulty of diagnosing active tuberculosis (TB) and lack of rapid drug susceptibility testing (DST) at the point of care remain critical obstacles to TB control. This report describes a high-intensity mycobacterium-specific-fluorophage (φ(2)GFP10) that for the first time allows direct visualization of Mycobacterium tuberculosis in clinical sputum samples. Engineered features distinguishing φ(2)GFP10 from previous reporter phages include an improved vector backbone with increased cloning capacity and superior expression of fluorescent reporter genes through use of an efficient phage promoter. φ(2)GFP10 produces a 100-fold increase in fluorescence per cell compared to existing reporter phages. DST for isoniazid and oxofloxacin, carried out in cultured samples, was complete within 36 h. Use of φ(2)GFP10 detected M. tuberculosis in clinical sputum samples collected from TB patients. DST for rifampin and kanamycin from sputum samples yielded results after 12 h of incubation with φ(2)GFP10. Fluorophage φ(2)GFP10 has potential for clinical development as a rapid, sensitive, and inexpensive point-of-care diagnostic tool for M. tuberculosis infection and for rapid DST.

Differences in Chlamydia trachomatis growth rates in human keratinocytes among lymphogranuloma venereum reference strains and clinical isolates.

The pathogenesis of the primary stage of lymphogranuloma venereum (LGV) is poorly understood. There is no skin cell model and LGV pathogenesis studies are therefore carried out on cells of different origin. Moreover, such studies usually use reference strains, which may have evolved over the years in culture. In this study, a model was developed in which Chlamydia trachomatis enters and grows in human keratinocytes at 37 and 33 °C. Keratinocytes were infected with fresh clinical isolates and the three LGV reference strains L1, L2 and L3. Growth was monitored for 5 days post-infection using fluorescence microscopy and image analysis software. Chlamydial replication was quicker at 37 than at 33 °C, despite 33 °C being the temperature of human skin. The serovar L2 reference strain grew significantly faster than the other strains, although the fresh clinical isolates were also serovar L2. When grown in keratinocytes at 33 °C, the L2 and L3 reference strains produced much larger inclusions than the other strains tested. This model, which utilizes keratinocytes, better simulates the conditions present at the initial site of infection in LGV than previously published literature, making it a useful tool for future LGV pathogenesis studies. In addition, the results indicate that fresh clinical isolates should be included in LGV pathogenesis studies.

Spread of extensively drug-resistant tuberculosis in KwaZulu-Natal province, South Africa.

BACKGROUND: In 2005 a cluster of 53 HIV-infected patients with extensively drug-resistant tuberculosis (XDR-TB) was detected in the Msinga sub-district, the catchment area for the Church of Scotland Hospital (CoSH) in Tugela Ferry, in KwaZulu-Natal province (KZN), South Africa. KZN is divided into 11 healthcare districts. We sought to determine the distribution of XDR TB cases in the province in relation to population density. METHODS: In this cross-sectional study, the KZN tuberculosis laboratory database was analysed. Results of all patients with a sputum culture positive for Mycobacterium tuberculosis from January 2006 to June 2007 were included. Drug-susceptibility test results for isoniazid, rifampicin, ethambutol, streptomycin, kanamycin and ofloxacin were available for all patients as well as the location of the hospital where their clinical diagnosis was made. FINDINGS: In total, 20858 patients attending one of 73 hospitals or their adjacent clinics had cultures positive for M. tuberculosis. Of these, 4170 (20%) were MDR-TB cases. Four hundred and forty three (11%) of the MDR tuberculosis cases displayed the XDR tuberculosis susceptibility profile. Only 1429 (34%) of the MDR-TB patients were seen at the provincial referral hospital for treatment. The proportion of XDR-TB amongst culture-confirmed cases was highest in the Msinga sub-district (19.6%), followed by the remaining part of the Umzinyati district (5.9%) and the other 10 districts (1.1%). The number of hospitals with at least one XDR-TB case increased from 18 (25%) to 58 (80%) during the study period. INTERPRETATION: XDR-TB is present throughout KZN. More than 65% of all diagnosed MDR-TB cases, including XDR-TB patients, were left untreated and likely remained in the community as a source of infection.

Increasing drug resistance in extensively drug-resistant tuberculosis, South Africa.

We expanded second-line tuberculosis (TB) drug susceptibility testing for extensively drug-resistant Mycobacterium tuberculosis isolates from South Africa. Of 19 patients with extensively drug-resistant TB identified during February 2008-April 2009, 13 (68%) had isolates resistant to all 8 drugs tested. This resistance leaves no effective treatment with available drugs in South Africa.

Rapid diagnosis of tuberculosis and multidrug resistance by the microscopic-observation drug-susceptibility assay.

RATIONALE: Mortality is exceedingly high and rapid among patients infected with HIV and tuberculosis (TB), in part because of limited access to appropriate TB diagnostics. The microscopic observation drug-susceptibility (MODS) assay is a simple, rapid, low-cost test for TB and multidrug-resistant (MDR) TB, but data in individuals infected with HIV and in Africa are limited. OBJECTIVES: To evaluate the MODS assay in a high-HIV-prevalence setting. METHODS: We performed a prospective diagnostic accuracy study of consecutive adults suspected to have TB from outpatient and inpatient settings at a district hospital in rural South Africa. Sputum was tested by concentrated smear microscopy; agar (Middlebrook 7H11) and liquid (mycobacterial growth indicator tube) culture; and the MODS assay. Drug-susceptibility testing (DST) was by indirect 1% proportion method and MODS. Reference standard for Mycobacterium tuberculosis detection was growth on Middlebrook or mycobacterial growth indicator tube culture; 1% proportion was the reference standard for isoniazid and rifampin DST. MEASUREMENTS AND MAIN RESULTS: Among 534 adults enrolled, 388 (73%) were HIV-positive, with a median CD4 count of 161 cells/mm(3) (interquartile range [IQR]: 72-307). TB was diagnosed by the reference standard culture in 113 (21%). MODS sensitivity was 85% (95% confidence interval [CI], 78-92%), and specificity was 97% (CI, 95-99%). MODS test performance did not differ by patients' HIV status (sensitivity 88% vs. 90%, specificity 97% vs. 100% for HIV-positive versus HIV-negative, respectively). For MDR-TB diagnosis (n = 11), sensitivity was 100% (one-sided CI, 68-100%) and specificity was 94% (CI, 82-98%). Median turnaround time for MDR-TB diagnosis was 7 days (IQR: 6-9) with MODS versus 70 days (IQR: 49-96) with indirect proportion method (P < 0.001). CONCLUSIONS: Among adult TB suspects predominantly infected with HIV, MODS provided high sensitivity and specificity for rapid diagnosis of TB and MDR-TB. Given the high mortality from TB and MDR-TB and prolonged opportunity for TB transmission before diagnosis, the MODS assay warrants serious consideration for use in similar high-HIV-prevalence, resource-limited settings.

Pyrido[1,2-a]benzimidazole-based agents active against tuberculosis (TB), multidrug-resistant (MDR) TB and extensively drug-resistant (XDR) TB.

The struggle against tuberculosis (TB) is still far from over. TB, caused by Mycobacterium tuberculosis, is one of the deadliest infections worldwide. Co-infection with human immunodeficiency virus (HIV) and the emergence of multidrug-resistant tuberculosis (MDR-TB) and extensively drug-resistant tuberculosis (XDR-TB) strains have further increased the burden for this disease. Herein, we report the discovery of 2-(4-chlorobenzyl)-3-methyl-1-oxo-1H,5H-pyrido[1,2-a]benzimidazole-4-carbonitrile as an effective antitubercular agent and the structural modifications of this molecule that have led to analogues with improved potency and lower toxicity. A number of these derivatives were also active at sub-micromolar concentrations against resistant TB strains and devoid of apparent toxicity to Vero cells, thereby underscoring their value as novel scaffolds for the development of new anti-TB drugs.

Einstein Contained Aerosol Pulmonizer (ECAP): Improved Biosafety for Multi-Drug Resistant (MDR) and Extensively Drug Resistant (XDR) Mycobacterium tuberculosis Aerosol Infection Studies.

A new apparatus enhances the biosafety of containment (biosafety level 3 [BSL-3]) and provides experimental reproducibility for aerosol infection experiments with MDR and XDR Mycobacterium tuberculosis. The methods are generally applicable to the study of airborne pathogens.