Clinical significance of Mycobacterium asiaticum isolates in Queensland, Australia

Mycobacterium asiaticum was first reported as a cause of human disease in 1982, with only a few cases in the literature to date. This study aims to review the clinical significance of M. asiaticum isolates in Queensland, Australia. A retrospective review (1989 to 2008) of patients with M. asiaticum isolates was conducted. Data were collected through the Queensland TB Control Centre database. Disease was defined in accordance with the American Thoracic Society criteria. Twenty-four patients (13 female) had a positive culture of M. asiaticum, many residing around the Tropic of Capricorn. M. asiaticum was responsible for pulmonary disease (n = 2), childhood lymphadenitis (n = 1), olecranon bursitis (n = 1), 6 cases of possible pulmonary disease, and 2 possible wound infections. Chronic lung disease was a risk factor for pulmonary infection, and wounds/lacerations were a risk factor for extrapulmonary disease. Extrapulmonary disease responded to local measures. Pulmonary disease responded to ethambutol-isoniazid-rifampin plus pyrazinamide for the first 2 months in one patient, and amikacin-azithromycin-minocycline in another patient. While M. asiaticum is rare in Queensland, there appears to be an environmental niche. Although often a colonizer, it can be a cause of pulmonary and extrapulmonary disease. Treatment of pulmonary disease remains challenging. Extrapulmonary disease does not mandate specific nontuberculous mycobacterium (NTM) treatment.

Evidence that human chlamydia pneumoniae was zoonotically acquired

Zoonotic infections are a growing threat to global health. Chlamydia pneumoniae is a major human pathogen that is widespread in human populations, causing acute respiratory disease, and has been associated with chronic disease. C. pneumoniae was first identified solely in human populations; however, its host range now includes other mammals, marsupials, amphibians, and reptiles. Australian koalas (Phascolarctos cinereus) are widely infected with two species of Chlamydia, C. pecorum and C. pneumoniae. Transmission of C. pneumoniae between animals and humans has not been reported; however, two other chlamydial species, C. psittaci and C. abortus, are known zoonotic pathogens. We have sequenced the 1,241,024-bp chromosome and a 7.5-kb cryptic chlamydial plasmid of the koala strain of C. pneumoniae (LPCoLN) using the whole-genome shotgun method. Comparative genomic analysis, including pseudogene and single-nucleotide polymorphism (SNP) distribution, and phylogenetic analysis of conserved genes and SNPs against the human isolates of C. pneumoniae show that the LPCoLN isolate is basal to human isolates. Thus, we propose based on compelling genomic and phylogenetic evidence that humans were originally infected zoonotically by an animal isolate(s) of C. pneumoniae which adapted to humans primarily through the processes of gene decay and plasmid loss, to the point where the animal reservoir is no longer required for transmission.

Kinematics of intracellular chlamydiae provide evidence for contact-dependent development

A crucial process of chlamydial development involves differentiation of the replicative reticulate body (RB) into the infectious elementary body (EB). We present experimental evidence to provide support for a contact-dependent hypothesis for explaining the trigger involved in differentiation. We recorded live-imaging of Chlamydia trachomatis-infected McCoy cells at key times during development and tracked the temporospatial trajectories of individual chlamydial particles. We found that movement of the particles is related to development. Early to mid-developmental stages involved slight wobbling of RBs. The average speed of particles increased sharply at 24 h postinfection (after the estimated onset of RB to EB differentiation). We also investigated a penicillin-supplemented culture containing EBs, RBs, and aberrantly enlarged, stressed chlamydiae. Near-immobile enlarged particles are consistent with their continued tethering to the chlamydial inclusion membrane (CIM). We found a significantly negative, nonlinear association between speed and size/type of particles, providing further support for the hypothesis that particles become untethered near the onset of RB to EB differentiation. This study establishes the relationship between the motion properties of the chlamydiae and developmental stages, whereby wobbling RBs gradually lose contact with the CIM, and RB detachment from the CIM is coincidental with the onset of late differentiation.

Mycobacterium lentiflavum in Drinking Water Supplies, Australia

Mycobacterium lentiflavum, a slow-growing nontuberculous mycobacterium, is a rare cause of human disease. It has been isolated from environmental samples worldwide. To assess the clinical significance of M. lentiflavum isolates reported to the Queensland Tuberculosis Control Centre, Australia, during 2001-2008, we explored the genotypic similarity and geographic relationship between isolates from humans and potable water in the Brisbane metropolitan area. A total of 47 isolates from 36 patients were reported; 4 patients had clinically significant disease. M. lentiflavum was cultured from 13 of 206 drinking water sites. These sites overlapped geographically with home addresses of the patients who had clinically significant disease. Automated repetitive sequence-based PCR genotyping showed a dominant environmental clone closely related to clinical strains. This finding suggests potable water as a possible source of M. lentiflavum infection in humans.

Heterogeneity of clinical and environmental isolates of Mycobacterium fortuitum using repetitive element sequence-based PCR: municipal water an unlikely source of community-acquired infections

M. fortuitum is a rapidly growing mycobacterium associated with community-acquired and nosocomial wound, soft tissue, and pulmonary infections. It has been postulated that water has been the source of infection especially in the hospital setting. The aim of this study was to determine if municipal water may be the source of community-acquired or nosocomial infections in the Brisbane area. Between 2007 and 2009, 20 strains of M. fortuitum were recovered from municipal water and 53 patients’ isolates were submitted to the reference laboratory. A wide variation in strain types was identified using repetitive element sequence-based PCR, with 13 clusters of ≥2 indistinguishable isolates, and 28 patterns consisting of individual isolates. The clusters could be grouped into seven similar groups (>95% similarity). Municipal water and clinical isolates collected during the same time period and from the same geographical area consisted of different strain types, making municipal water an unlikely source of sporadic human infection.

Strain variation amongst clinical and potable water isolates of M.kansasii using automated repetitive unit PCR

Mycobacterium kansasii is a pulmonary pathogen that has been grown readily from municipal water, but rarely isolated from natural waters. A definitive link between water exposure and disease has not been demonstrated and the environmental niche for this organism is poorly understood. Strain typing of clinical isolates has revealed seven subtypes with Type 1 being highly clonal and responsible for most infections worldwide. The prevalence of other subtypes varies geographically. In this study 49 water isolates are compared with 72 patient isolates from the same geographical area (Brisbane, Australia), using automated repetitive unit PCR (Diversilab) and ITS RFLP. The clonality of the dominant clinical strain type is again demonstrated but with rep-PCR, strain variation within this group is evident comparable with other reported methods. There is significant heterogeneity of water isolates and very few are similar or related to the clinical isolates. This suggests that if water or aerosol transmission is the mode of infection, then point source contamination likely occurs from an alternative environmental source.

Antibodies mediate formation of neutrophil extracellular traps in the middle ear and facilitate secondary pneumococcal otitis media

Otitis media (OM) (a middle ear infection) is a common childhood illness that can leave some children with permanent hearing loss. OM can arise following infection with a variety of different pathogens, including a coinfection with influenza A virus (IAV) and Streptococcus pneumoniae (the pneumococcus). We and others have demonstrated that coinfection with IAV facilitates the replication of pneumococci in the middle ear. Specifically, we used a mouse model of OM to show that IAV facilitates the outgrowth of S. pneumoniae in the middle ear by inducing middle ear inflammation. Here, we seek to understand how the host inflammatory response facilitates bacterial outgrowth in the middle ear. Using B cell-deficient infant mice, we show that antibodies play a crucial role in facilitating pneumococcal replication. We subsequently show that this is due to antibody-dependent neutrophil extracellular trap (NET) formation in the middle ear, which, instead of clearing the infection, allows the bacteria to replicate. We further demonstrate the importance of these NETs as a potential therapeutic target through the transtympanic administration of a DNase, which effectively reduces the bacterial load in the middle ear. Taken together, these data provide novel insight into how pneumococci are able to replicate in the middle ear cavity and induce disease.

Genome-wide mapping of cystitis due to Streptococcus agalactiae and Escherichia coli in mice identifies a unique bladder transcriptome that signifies pathogen-specific antimicrobial defense against urinary tract infection

The most common causes of urinary tract infections (UTIs) are Gram-negative pathogens such as Escherichia coli; however, Gram-positive organisms including Streptococcus agalactiae, or group B streptococcus (GBS), also cause UTI. In GBS infection, UTI progresses to cystitis once the bacteria colonize bladder, but the host responses triggered in the bladder immediately following infection are largely unknown. Here, we used genome-wide expression profiling to map the bladder transcriptome of GBS UTI in mice infected transurethrally with uropathogenic GBS that was cultured from a 35 year-old women with cystitis. RNA from bladders was applied to Affymetrix Gene-1.0ST microarrays; qRT-PCR was used to analyze selected gene responses identified in array datasets. A surprisingly small significant gene list of 172 genes was identified at 24h; this compared to 2507 genes identified in a side-by-side comparison with uropathogenic E. coli (UPEC). No genes exhibited significantly altered expression at 2h in GBS-infected mice according to arrays despite high bladder bacterial loads at this early time point. The absence of a marked early host response to GBS juxtaposed with broad-based bladder responses activated by UPEC at 2h. Bioinformatics analyses including integrative systems-level network mapping revealed multiple activated biological pathways in the GBS cystitis transcriptome that regulate leukocyte activation, inflammation, apoptosis, and cytokine-chemokine biosynthesis. These findings define a novel, minimalistic type of bladder host response triggered by GBS UTI, which comprises collective antimicrobial pathways that differ dramatically from those activated by UPEC. Overall, this study emphasizes the unique nature of bladder immune activation mechanisms triggered by distinct uropathogens.

Current understanding of streptococcal urinary tract infection

Group B streptococcus (GBS), also known as Streptococcus agalactiae is a Gram-positive, β-hemolytic, chain-forming bacterium and a commensal within the genital tract flora in approximately 25% of healthy adult women (Campbell et al., 2000). The organism is a leading cause of serious infection in newborns, pregnant women, and older persons with chronic medical illness (Baker et al., Edwards&Baker, 2005). In neonates GBS infection most commonly causes pneumonia, meningitis, and sepsis. In addition to maternal cervicovaginal colonization and neonatal infection that can result from vertical transmission of GBS from mothers to their infants, the bacterium can also cause urinary tract infection (UTI). The spectrum of GBS UTI includes asymptomatic bacteriuria (ABU), cystitis, pyelonephritis, urethritis, and urosepsis (Bronsema et al., 1993, Edwards&Baker, 2005, Farley et al., 1993, Lefevre et al., 1991, McKenna et al., 2003, Munoz et al., 1992, Ulett et al., 2009). GBS ABU is particularly common among pregnant women, although those most at risk for cystitis due to GBS appear to be elderly individuals (Edwards&Baker, 2005, Falagas et al., 2006, Muller et al., 2006). In addition to acute and asymptomatic UTI other invasive diseases caused by GBS infection include skin infections, bacteraemia, pneumonia, arthritis, and endocarditis (Liston et al., 1979, Patil & Martin, 2010, Tissi et al., 1997, Trivalle et al., 1998). Thus, GBS is considered unique in terms of its ability to cause a spectrum of diseases in newborns and adult humans and its ability to colonize the genital tract of healthy women in a commensal-type manner...

Pedagogical tools designed to promote engagement and active learning in the biomedical sciences and allied health disciplines

Perceiving students, science students especially, as mere consumers of facts and information belies the importance of a need to engage them with the principles underlying those facts and is counter-intuitive to the facilitation of knowledge and understanding. Traditional didactic lecture approaches need a re-think if student classroom engagement and active learning are to be valued over fact memorisation and fact recall. In our undergraduate biomedical science programs across Years 1, 2 and 3 in the Faculty of Health at QUT, we have developed an authentic learning model with an embedded suite of pedagogical strategies that foster classroom engagement and allow for active learning in the sub-discipline area of medical bacteriology. The suite of pedagogical tools we have developed have been designed to enable their translation, with appropriate fine-tuning, to most biomedical and allied health discipline teaching and learning contexts. Indeed, aspects of the pedagogy have been successfully translated to the nursing microbiology study stream at QUT. The aims underpinning the pedagogy are for our students to: (1) Connect scientific theory with scientific practice in a more direct and authentic way, (2) Construct factual knowledge and facilitate a deeper understanding, and (3) Develop and refine their higher order flexible thinking and problem solving skills, both semi-independently and independently. The mindset and role of the teaching staff is critical to this approach since for the strategy to be successful tertiary teachers need to abandon traditional instructional modalities based on one-way information delivery. Face-to-face classroom interactions between students and lecturer enable realisation of pedagogical aims (1), (2) and (3). The strategy we have adopted encourages teachers to view themselves more as expert guides in what is very much a student-focused process of scientific exploration and learning. Specific pedagogical strategies embedded in the authentic learning model we have developed include: (i) interactive lecture-tutorial hybrids or lectorials featuring teacher role-plays as well as class-level question-and-answer sessions, (ii) inclusion of “dry” laboratory activities during lectorials to prepare students for the wet laboratory to follow, (iii) real-world problem-solving exercises conducted during both lectorials and wet laboratory sessions, and (iv) designing class activities and formative assessments that probe a student’s higher order flexible thinking skills. Flexible thinking in this context encompasses analytical, critical, deductive, scientific and professional thinking modes. The strategic approach outlined above is designed to provide multiple opportunities for students to apply principles flexibly according to a given situation or context, to adapt methods of inquiry strategically, to go beyond mechanical application of formulaic approaches, and to as much as possible self-appraise their own thinking and problem solving. The pedagogical tools have been developed within both workplace (real world) and theoretical frameworks. The philosophical core of the pedagogy is a coherent pathway of teaching and learning which we, and many of our students, believe is more conducive to student engagement and active learning in the classroom. Qualitative and quantitative data derived from online and hardcopy evaluations, solicited and unsolicited student and graduate feedback, anecdotal evidence as well as peer review indicate that: (i) our students are engaging with the pedagogy, (ii) a constructivist, authentic-learning approach promotes active learning, and (iii) students are better prepared for workplace transition.