Concentration of Lymph Node Aspirate Improves the Sensitivity of Acid Fast Smear Microscopy for the Diagnosis of Tuberculous Lymphadenitis in Jimma,Southwest Ethiopia

Background

Tuberculous lymphadenitis (TBLN) is the most common form of extrapulmonary tuberculosis. The cytomorphological features of lymph node smears have reduced specificity for the diagnosis of tuberculosis. The diagnosis of TBLN with direct smear microscopy lacks sensitivity due to the limited number of bacilli in lymph node aspirate. Therefore, we aimed to assess whether the concentration of lymph node aspirate improves the sensitivity of acid fast smear microscopy for the diagnosis of tuberculous lymphadenitis.

Methods

A cross-sectional comparative study was conducted on 200 patients clinically suspected for tuberculous lymphadenitis in Jimma, Ethiopia. Lymph node aspirate was collected. The first two drops were used for cytomorphological study and direct acid fast staining. The remaining aspirate was treated with N-acetyl-L-cysteine (NALC) and concentrated by centrifugation at 3000 g for 15 minutes. The sediment was used for acid fast staining and culture. Differentiation of M. tuberculosis complex (MTBC) from non-tuberculous mycobacteria (NTM) was done by para-nitrobenzoic acid susceptibility test.

Result

Complete data were available for 187 study subjects. 68% (127/187) were positive for M. tuberculosis on culture. Four isolates, 2.1% (4/187), were identified as NTM. The detection rate of direct smear microscopy was 25.1% and that of the concentration method 49.7%. Cytomorphologically, 79.7% of cases were classified as TBLN. The sensitivity of direct smear microscopy was 34.6%, for concentrated smear microscopy 66.1%, and for cytomorphology 89.8%. Two AFB positive cases on concentration method were non-tuberculosis mycobacteria (NTM). The concentration method yielded a positive result from seven cases diagnosed as suppurative abscess by cytology. Both for the direct and concentration methods the highest rate of AFB positivity was observed in smears showing caseous necrosis alone. Smear positivity rate decreased with the appearance of epithelioid cell aggregates.

Conclusion

The concentration of lymph node aspirates for acid fast smear microscopy had significantly higher sensitivity than direct microscopy.     

Invasive Pneumococcal Isolates from Danish Infants (0 - 90 Days) during the Years 1943 to 2013

Background

The seven-valent pneumococcal conjugate vaccine (PCV-7) was introduced in the Danish childhood immunization program (at 3, 5 and 12 months of age) in 2007 and was replaced with PCV-13 in 2010 without changes to the schedule. After the introduction of these vaccines the incidence of invasive pneumococcal disease (IPD) due to vaccine types (VTs) declined markedly in children aged 0–2 years; however, cases among infants too young to be protected by vaccination have not been studied in detail. We present data on IPD in infants less than 90 days from 1943 until 2013.

Study design

The study included all infants younger than 90 days born from 1943 through 2013, who had not been PCV vaccinated and from whom a pneumococcus isolate from blood or cerebrospinal fluid had been submitted to the Danish national reference laboratory. All isolates were serotyped using Pneumotest Latex and Quellung reaction.

Results

A total of 216 IPD cases were identified. The age group specific incidence (total number of IPD cases per 100,000 live births) varied from 0 to 16 in the period 1943 to 2007 and from 1.7 to 9.2 in the period 2008 to 2013. IPD cases due to PCV-7 serotypes were not observed later than 2009.

Conclusion

In Danish infants younger than 90 days, IPD due to PCV-7 serotypes has decreased and has not been observed since 2009, but the total incidence of IPD has not changed.     

Genome Sequencing and Analysis of BCG Vaccine Strains

Background

Although the Bacillus Calmette-Guérin (BCG) vaccine against tuberculosis (TB) has been available for more than 75 years, one third of the world''s population is still infected with Mycobacterium tuberculosis and approximately 2 million people die of TB every year. To reduce this immense TB burden, a clearer understanding of the functional genes underlying the action of BCG and the development of new vaccines are urgently needed.

Methods and Findings

Comparative genomic analysis of 19 M. tuberculosis complex strains showed that BCG strains underwent repeated human manipulation, had higher region of deletion rates than those of natural M. tuberculosis strains, and lost several essential components such as T-cell epitopes. A total of 188 BCG strain T-cell epitopes were lost to various degrees. The non-virulent BCG Tokyo strain, which has the largest number of T-cell epitopes (359), lost 124. Here we propose that BCG strain protection variability results from different epitopes. This study is the first to present BCG as a model organism for genetics research. BCG strains have a very well-documented history and now detailed genome information. Genome comparison revealed the selection process of BCG strains under human manipulation (1908–1966).

Conclusions

Our results revealed the cause of BCG vaccine strain protection variability at the genome level and supported the hypothesis that the restoration of lost BCG Tokyo epitopes is a useful future vaccine development strategy. Furthermore, these detailed BCG vaccine genome investigation results will be useful in microbial genetics, microbial engineering and other research fields.     

Molecular Characterization of Invasive Meningococcal Isolates from Countries in the African Meningitis Belt before Introduction of a Serogroup A Conjugate Vaccine

Background

The serogroup A conjugate meningococcal vaccine, MenAfriVac, was introduced in mass vaccination campaigns in December 2010 in Burkina Faso, Mali and Niger. In the coming years, vaccination will be extended to other African countries at risk of epidemics. To document the molecular characteristics of disease-causing meningococcal strains circulating in the meningitis belt of Africa before vaccine introduction, the World Health Organization Collaborating Centers on Meningococci in Europe and United States established a common strain collection of 773 isolates from cases of invasive meningococcal disease collected between 2004 and 2010 from 13 sub-Saharan countries.

Methodology

All isolates were characterized by multilocus sequence typing, and 487 (62%) were also analyzed for genetic variation in the surface antigens PorA and FetA. Antibiotic susceptibility was tested for part of the collection.

Principal Findings

Only 19 sequence types (STs) belonging to 6 clonal complexes were revealed. ST-5 clonal complex dominated with 578 (74.8%) isolates. All ST-5 complex isolates were remarkably homogeneous in their PorA (P1.20,9) and FetA (F3-1) and characterized the serogroup A strains which have been responsible for most epidemics during this time period. Sixty-eight (8.8%) of the 773 isolates belonged to the ST-11 clonal complex which was mainly represented by serogroup W135, while an additional 38 (4.9%) W135 isolates belonged to the ST-175 complex. Forty-eight (6.2%) serogroup X isolates from West Africa belonged to the ST-181 complex, while serogroup X cases in Kenya and Uganda were caused by an unrelated clone, ST-5403. Serogroup X, ST-181, emerged in Burkina Faso before vaccine introduction.

Conclusions

In the seven years preceding introduction of a new serogroup A conjugate vaccine, serogroup A of the ST-5 clonal complex was identified as the predominant disease-causing strain.     

Protective efficacy of BCG overexpressing an L,D-transpeptidase against M. tuberculosis infection

Background

M. bovis Bacille Calmette-Guérin (BCG), currently the only available vaccine against tuberculosis (TB), fails to adequately protect individuals from active and latent TB infection. New vaccines are desperately needed to decrease the worldwide burden of TB.

Methods and Findings

We created a recombinant strain of BCG that overproduces an L,D-transpeptidase in order to alter the bacterial peptidoglycan layer and consequently increase the ability of this immunogen to protect against virulent M. tuberculosis (Mtb). We demonstrate that this novel recombinant BCG protects mice against virulent Mtb at least as well as control BCG, as measured by its ability to reduce bacterial burden in lungs and spleen, reduce lung histopathology, and prolong survival. A nutrient starved recombinant BCG preparation, while offering comparable protection, elicited a response characterized by elevated levels of select Th1 cytokines.

Conclusions

Recombinant BCG overexpressing a L,D-transpeptidase that is nutrient starved elicits a stronger Th1 type response and is at least as protective as parent BCG. Results from this study suggest that nutrient starvation treatment of live BCG vaccines should be further investigated as a way to increase host induction of Th-1 related cytokines in the development of experimental anti-TB vaccines.     

The long-term effect of current and new interventions on the new case detection of leprosy: a modeling study

Background

Although the number of newly detected leprosy cases has decreased globally, a quarter of a million new cases are detected annually and eradication remains far away. Current options for leprosy prevention are contact tracing and BCG vaccination of infants. Future options may include chemoprophylaxis and early diagnosis of subclinical infections. This study compared the predicted trends in leprosy case detection of future intervention strategies.

Methods

Seven leprosy intervention scenarios were investigated with a microsimulation model (SIMCOLEP) to predict future leprosy trends. The baseline scenario consisted of passive case detection, multidrug therapy, contact tracing, and BCG vaccination of infants. The other six scenarios were modifications of the baseline, as follows: no contact tracing; with chemoprophylaxis; with early diagnosis of subclinical infections; replacement of the BCG vaccine with a new tuberculosis vaccine ineffective against Mycobacterium leprae (“no BCG”); no BCG with chemoprophylaxis; and no BCG with early diagnosis.

Findings

Without contact tracing, the model predicted an initial drop in the new case detection rate due to a delay in detecting clinical cases among contacts. Eventually, this scenario would lead to new case detection rates higher than the baseline program. Both chemoprophylaxis and early diagnosis would prevent new cases due to a reduction of the infectious period of subclinical cases by detection and cure of these cases. Also, replacing BCG would increase the new case detection rate of leprosy, but this effect could be offset with either chemoprophylaxis or early diagnosis.

Conclusions

This study showed that the leprosy incidence would be reduced substantially by good BCG vaccine coverage and the combined strategies of contact tracing, early diagnosis, and treatment of infection and/or chemoprophylaxis among household contacts. To effectively interrupt the transmission of M. leprae, it is crucial to continue developing immuno- and chemoprophylaxis strategies and an effective test for diagnosing subclinical infections.     

Comparative tuberculosis (TB) prevention effectiveness in children of Bacillus Calmette-Guérin (BCG) vaccines from different sources, Kazakhstan

Background

Except during a 1-year period when BCG vaccine was not routinely administered, annual coverage of infants with Bacillus Calmette-Guérin (BCG) in Kazakhstan since 2002 has exceeded 95%. BCG preparations from different sources (Japan, Serbia, and Russia) or none were used exclusively in comparable 7-month time-frames, September through March, in 4 successive years beginning in 2002. Our objective was to assess relative effectiveness of BCG immunization.

Methods/Findings

We compared outcomes of birth cohorts from the 4 time-frames retrospectively. Three cohorts received vaccine from one of three manufacturers exclusively, and one cohort was not vaccinated. Cohorts were followed for 3 years for notifications of clinical TB and of culture-confirmed TB, and for 21 months for TB meningitis notifications. Prevention effectiveness based on relative risk of TB incidence was calculated for each vaccinated cohort compared to the non-vaccinated cohort.Although there were differences in prevention effectiveness observed among the three BCG vaccines, all were protective. The Japanese vaccine (currently used in Kazakhstan), the Serbian vaccine, and the Russian vaccine respectively were 69%, 43%, and 22% effective with respect to clinical TB notifications, and 92%, 82%, and 51% effective with respect to culture confirmed TB. All three vaccines were >70% effective with respect to TB meningitis.

Limitations

Potential limitations included considerations that 1) the methodology used was retrospective, 2) multiple risk factors could have varied between cohorts and affected prevention effectiveness measures, 3) most cases were clinically diagnosed, and TB culture-positive case numbers and TB meningitis case numbers were sparse, and 4) small variations in reported population TB burden could have affected relative risk of exposure for cohorts.

Conclusions/Significance

All three BCG vaccines evaluated were protective against TB, and prevention effectiveness varied by manufacturer. When setting national immunization policy, consideration should be given to prevention effectiveness of BCG preparations.     

Reducing the Activity and Secretion of Microbial Antioxidants Enhances the Immunogenicity of BCG

Background

In early clinical studies, the live tuberculosis vaccine Mycobacterium bovis BCG exhibited 80% protective efficacy against pulmonary tuberculosis (TB). Although BCG still exhibits reliable protection against TB meningitis and miliary TB in early childhood it has become less reliable in protecting against pulmonary TB. During decades of in vitro cultivation BCG not only lost some genes due to deletions of regions of the chromosome but also underwent gene duplication and other mutations resulting in increased antioxidant production.

Methodology/Principal Findings

To determine whether microbial antioxidants influence vaccine immunogenicity, we eliminated duplicated alleles encoding the oxidative stress sigma factor SigH in BCG Tice and reduced the activity and secretion of iron co-factored superoxide dismutase. We then used assays of gene expression and flow cytometry with intracellular cytokine staining to compare BCG-specific immune responses in mice after vaccination with BCG Tice or the modified BCG vaccine. Compared to BCG, the modified vaccine induced greater IL-12p40, RANTES, and IL-21 mRNA in the spleens of mice at three days post-immunization, more cytokine-producing CD8+ lymphocytes at the peak of the primary immune response, and more IL-2-producing CD4+ lymphocytes during the memory phase. The modified vaccine also induced stronger secondary CD4+ lymphocyte responses and greater clearance of challenge bacilli.

Conclusions/Significance

We conclude that antioxidants produced by BCG suppress host immune responses. These findings challenge the hypothesis that the failure of extensively cultivated BCG vaccines to prevent pulmonary tuberculosis is due to over-attenuation and suggest instead a new model in which BCG evolved to produce more immunity-suppressing antioxidants. By targeting these antioxidants it may be possible to restore BCG''s ability to protect against pulmonary TB.     

Effectiveness of Routine BCG Vaccination on Buruli Ulcer Disease: A Case-Control Study in the Democratic Republic of Congo,Ghana and Togo

Background

The only available vaccine that could be potentially beneficial against mycobacterial diseases contains live attenuated bovine tuberculosis bacillus (Mycobacterium bovis) also called Bacillus Calmette-Guérin (BCG). Even though the BCG vaccine is still widely used, results on its effectiveness in preventing mycobacterial diseases are partially contradictory, especially regarding Buruli Ulcer Disease (BUD). The aim of this case-control study is to evaluate the possible protective effect of BCG vaccination on BUD.

Methodology

The present study was performed in three different countries and sites where BUD is endemic: in the Democratic Republic of the Congo, Ghana, and Togo from 2010 through 2013. The large study population was comprised of 401 cases with laboratory confirmed BUD and 826 controls, mostly family members or neighbors.

Principal Findings

After stratification by the three countries, two sexes and four age groups, no significant correlation was found between the presence of BCG scar and BUD status of individuals. Multivariate analysis has shown that the independent variables country (p = 0.31), sex (p = 0.24), age (p = 0.96), and presence of a BCG scar (p = 0.07) did not significantly influence the development of BUD category I or category II/III. Furthermore, the status of BCG vaccination was also not significantly related to duration of BUD or time to healing of lesions.

Conclusions

In our study, we did not observe significant evidence of a protective effect of routine BCG vaccination on the risk of developing either BUD or severe forms of BUD. Since accurate data on BCG strains used in these three countries were not available, no final conclusion can be drawn on the effectiveness of BCG strain in protecting against BUD. As has been suggested for tuberculosis and leprosy, well-designed prospective studies on different existing BCG vaccine strains are needed also for BUD.     

Cellular immune responses to nine Mycobacterium tuberculosis vaccine candidates following intranasal vaccination

Background

The identification of Mycobacterium tuberculosis vaccines that elicit a protective immune response in the lungs is important for the development of an effective vaccine against tuberculosis.

Methods and Principal Findings

In this study, a comparison of intranasal (i.n.) and subcutaneous (s.c.) vaccination with the BCG vaccine demonstrated that a single moderate dose delivered intranasally induced a stronger and sustained M. tuberculosis-specific T-cell response in lung parenchyma and cervical lymph nodes of BALB/c mice than vaccine delivered subcutaneously. Both BCG and a multicomponent subunit vaccine composed of nine M. tuberculosis recombinant proteins induced strong antigen-specific T-cell responses in various local and peripheral immune compartments. Among the nine recombinant proteins evaluated, the alanine proline rich antigen (Apa, Rv1860) was highly antigenic following i.n. BCG and immunogenic after vaccination with a combination of the nine recombinant antigens. The Apa-induced responses included induction of both type 1 and type 2 cytokines in the lungs as evaluated by ELISPOT and a multiplexed microsphere-based cytokine immunoassay. Of importance, i.n. subunit vaccination with Apa imparted significant protection in the lungs and spleen of mice against M. tuberculosis challenge. Despite observed differences in the frequencies and location of specific cytokine secreting T cells both BCG vaccination routes afforded comparable levels of protection in our study.

Conclusion and Significance

Overall, our findings support consideration and further evaluation of an intranasally targeted Apa-based vaccine to prevent tuberculosis.     

Intranasal Mucosal Boosting with an Adenovirus-Vectored Vaccine Markedly Enhances the Protection of BCG-Primed Guinea Pigs against Pulmonary Tuberculosis

Background

Recombinant adenovirus-vectored (Ad) tuberculosis (TB) vaccine platform has demonstrated great potential to be used either as a stand-alone or a boost vaccine in murine models. However, Ad TB vaccine remains to be evaluated in a more relevant and sensitive guinea pig model of pulmonary TB. Many vaccine candidates shown to be effective in murine models have subsequently failed to pass the test in guinea pig models.

Methods and Findings

Specific pathogen-free guinea pigs were immunized with BCG, AdAg85A intranasally (i.n), AdAg85A intramuscularly (i.m), BCG boosted with AdAg85A i.n, BCG boosted with AdAg85A i.m, or treated only with saline. The animals were then infected by a low-dose aerosol of M. tuberculosis (M.tb). At the specified times, the animals were sacrificed and the levels of infection in the lung and spleen were assessed. In separate studies, the long-term disease outcome of infected animals was monitored until the termination of this study. Immunization with Ad vaccine alone had minimal beneficial effects. Immunization with BCG alone and BCG prime-Ad vaccine boost regimens significantly reduced the level of M.tb infection in the tissues to a similar extent. However, while BCG alone prolonged the survival of infected guinea pigs, the majority of BCG-immunized animals succumbed by 53 weeks post-M.tb challenge. In contrast, intranasal or intramuscular Ad vaccine boosting of BCG-primed animals markedly improved the survival rate with 60% of BCG/Ad i.n- and 40% of BCG/Ad i.m-immunized guinea pigs still surviving by 74 weeks post-aerosol challenge.

Conclusions

Boosting, particularly via the intranasal mucosal route, with AdAg85A vaccine is able to significantly enhance the long-term survival of BCG-primed guinea pigs following pulmonary M.tb challenge. Our results thus support further evaluation of this viral-vectored TB vaccine in clinical trials.     

HIV Case Notification Rates in the Kingdom of Saudi Arabia over the Past Decade (2000–2009)

Objective

To study trends in HIV case notification rates in the Kingdom of Saudi Arabia.

Design

A ten year retrospective review of annual HIV case notification returns to the Ministry of Health, Kingdom of Saudi Arabia.

Methods

Annual Registry statistics covering the period 2000 to 2009 were reviewed. Annual incidence trends were stratified according to the following demographics: age, nationality, geographical region of residence, gender, and mode of disease acquisition.

Results

10,217 new HIV cases (2,956 in Saudi nationals and 7,261 in non-Saudis) were reported. Africans of Sub-Saharan Africa origin accounting for 3,982/7,261 (53%) of non-Saudi cases constituted: Ethiopians (2,271), Nigerians (1,048), and Sudanese nationals (663). The overall average annual incidence was <4 cases per 100,000; 1.5 cases per 100,000 for Saudis (range 0.5–2.5), and 13.2 per 100,000 for non-Saudis (range 5.7–19.0). Notifications increased yearly from 2000 for both groups until a plateau was reached in 2006 at 1,390 new cases. Case notification in Saudi nationals increased from 20% in the early 2001 to 40% in 2009. 4% (124/2,956) of cases were reported in Saudi children. The male to female ratio was 1.6∶1 for non-Saudi nationals (43.8% male, 27.3% female) and 4.4∶1 for Saudis (23.5% male, 5.4% female).

Conclusions

Whilst the numbers of reported HIV cases have stabilised since 2006, HIV/AIDS remains an important public health problem in KSA, both in migrants and Saudi nationals. HIV transmission to Saudi children is also of concern. Optimization of data collection, surveillance, and pro-active screening for HIV is required.     

Safety and immunogenicity of the candidate tuberculosis vaccine MVA85A in West Africa

Background

Vaccination with a recombinant modified vaccinia Ankara expressing antigen 85A from Mycobacterium tuberculosis, MVA85A, induces high levels of cellular immune responses in UK volunteers. We assessed the safety and immunogenicity of this new vaccine in West African volunteers.

Methods and Findings

We vaccinated 21 healthy adult male subjects (11 BCG scar negative and 10 BCG scar positive) with MVA85A after screening for evidence of prior exposure to mycobacteria. We monitored them over six months, observing for clinical, haematological and biochemical adverse events, together with assessment of the vaccine induced cellular immune response using ELISPOT and flow cytometry. MVA85A was well tolerated with no significant adverse events. Mild local and systemic adverse events were consistent with previous UK trials. Marked immunogenicity was found whether individuals had a previous BCG scar or not. There was not enhanced immunogenicity in those with a BCG scar, and induced T cell responses were better maintained in apparently BCG-naïve Gambians than previously studied BCG-naïve UK vaccinees. Although responses were predominantly attributable to CD4+ T cells, we also identified antigen specific CD8+ T cell responses, in subjects who were HLA B-35 and in whom enough blood was available for more detailed immunological analysis.

Conclusions

These data on the safety and immunogenicity of MVA85A in West Africa support its accelerated development as a promising booster vaccine for tuberculosis.

Trial Registration

ClinicalTrials.gov {"type":"clinical-trial","attrs":{"text":"NCT00423839","term_id":"NCT00423839"}}NCT00423839     

PspA Family Distribution,Antimicrobial Resistance and Serotype of Streptococcus pneumoniae Isolated from Upper Respiratory Tract Infections in Japan

Background

The protection against pneumococcal infections provided by currently available pneumococcal polysaccharide conjugate vaccines are restricted to the limited number of the serotypes included in the vaccine. In the present study, we evaluated the distribution of the pneumococcal capsular type and surface protein A (PspA) family of pneumococcal isolates from upper respiratory tract infections in Japan.

Methods

A total of 251 S. pneumoniae isolates from patients seeking treatment for upper respiratory tract infections were characterized for PspA family, antibiotic resistance and capsular type.

Results

Among the 251 pneumococci studied, the majority (49.4%) was identified as belonging to PspA family 2, while most of the remaining isolates (44.6%) belonged to family 1. There were no significant differences between the distributions of PspA1 versus PspA2 isolates based on the age or gender of the patient, source of the isolates or the isolates’ susceptibilities to penicillin G. In contrast, the frequency of the mefA gene presence and of serotypes 15B and 19F were statistically more common among PspA2 strains.

Conclusion

The vast majority of pneumococci isolated from the middle ear fluids, nasal discharges/sinus aspirates or pharyngeal secretions represented PspA families 1 and 2. Capsular serotypes were generally not exclusively associated with certain PspA families, although some capsular types showed a much higher proportion of either PspA1 or PspA2. A PspA-containing vaccine would potentially provide high coverage against pneumococcal infectious diseases because it would be cross-protective versus invasive disease with the majority of pneumococci infecting children and adults.     

Determining Vaccination Frequency in Farmed Rainbow Trout Using Vibrio anguillarum O1 Specific Serum Antibody Measurements

Background

Despite vaccination with a commercial vaccine with a documented protective effect against Vibrio anguillarum O1 disease outbreaks caused by this bacterium have been registered among rainbow trout at Danish fish farms. The present study examined specific serum antibody levels as a valid marker for assessing vaccination status in a fish population. For this purpose a highly sensitive enzyme-linked immunosorbent assay (ELISA) was developed and used to evaluate sera from farmed rainbow trout vaccinated against V. anguillarum O1.

Study Design

Immune sera from rainbow trout immunised with an experimental vaccine based on inactivated V. anguillarum O1 bacterin in Freund’s incomplete adjuvant were used for ELISA optimisation. Subsequently, sera from farmed rainbow trout vaccinated with a commercial vaccine against V. anguillarum were analysed with the ELISA. The measured serum antibody levels were compared with the vaccine status of the fish (vaccinated/unvaccinated) as evaluated through visual examination.

Results

Repeated immunisation with the experimental vaccine lead to increasing levels of specific serum antibodies in the vaccinated rainbow trout. The farmed rainbow trout responded with high antibody levels to a single injection with the commercial vaccine. However, the diversity in responses was more pronounced in the farmed fish. Primary visual examinations for vaccine status in rainbow trout from the commercial farm revealed a large pool of unvaccinated specimens (vaccination failure rate = 20%) among the otherwise vaccinated fish. Through serum analyses using the ELISA in a blinded set-up it was possible to separate samples collected from the farmed rainbow trout into vaccinated and unvaccinated fish.

Conclusions

Much attention has been devoted to development of new and more effective vaccines. Here we present a case from a Danish rainbow trout farm indicating that attention should also be directed to the vaccination procedure in order to secure high vaccination frequencies necessary for optimal protection with a reported effective vaccine.     

Genetic Characterisation of Malawian Pneumococci Prior to the Roll-Out of the PCV13 Vaccine Using a High-Throughput Whole Genome Sequencing Approach

Background

Malawi commenced the introduction of the 13-valent pneumococcal conjugate vaccine (PCV13) into the routine infant immunisation schedule in November 2011. Here we have tested the utility of high throughput whole genome sequencing to provide a high-resolution view of pre-vaccine pneumococcal epidemiology and population evolutionary trends to predict potential future change in population structure post introduction.

Methods

One hundred and twenty seven (127) archived pneumococcal isolates from randomly selected adults and children presenting to the Queen Elizabeth Central Hospital, Blantyre, Malawi underwent whole genome sequencing.

Results

The pneumococcal population was dominated by serotype 1 (20.5% of invasive isolates) prior to vaccine introduction. PCV13 is likely to protect against 62.9% of all circulating invasive pneumococci (78.3% in under-5-year-olds). Several Pneumococcal Molecular Epidemiology Network (PMEN) clones are now in circulation in Malawi which were previously undetected but the pandemic multidrug resistant PMEN1 lineage was not identified. Genome analysis identified a number of novel sequence types and serotype switching.

Conclusions

High throughput genome sequencing is now feasible and has the capacity to simultaneously elucidate serotype, sequence type and as well as detailed genetic information. It enables population level characterization, providing a detailed picture of population structure and genome evolution relevant to disease control. Post-vaccine introduction surveillance supported by genome sequencing is essential to providing a comprehensive picture of the impact of PCV13 on pneumococcal population structure and informing future public health interventions.     

The Feasibility of the Interferon Gamma Release Assay and Predictors of Discordance with the Tuberculin Skin Test for the Diagnosis of Latent Tuberculosis Infection in a Remote Aboriginal Community

Background

The tuberculin skin test (TST) is the standard test used to screen for latent TB infection (LTBI) in the northern Canadian territory of Nunavut. Interferon gamma release assays (IGRA) are T cell blood-based assays to diagnose LTBI. The Bacillus Calmette-Guerin (BCG) vaccine is part of the routine immunization schedule in Nunavut. The objective of this study was to test the feasibility, and predictors of discordance between the Tuberculin Skin Test (TST) and the IGRA assay in a medically under-serviced remote arctic Aboriginal population.

Methods

Both the TST and QuantiFERON-TB Gold (Qiagen group) IGRA tests were offered to people in their homes as part of a public health campaign aimed at high TB risk residential areas in Iqaluit, Nunavut, Canada. Feasibility was measured by the capacity of the staff to do the test successfully as measured by the proportion of results obtained.

Results

In this population of predominantly young Inuit who were mostly BCG vaccinated, the use of IGRA for the diagnosis of LTBI was feasible. IGRA testing resulted in more available test results reaching patients (95.6% vs 90.9% p = 0.02) but took longer (median 8 days (IGRA) vs 2 days (TST), p value <0.0001). 44/256 participants (17.2%) had discordant results. Multivariable regression analysis suggested that discordant results were most likely to have received multiple BCG vaccinations (RR 20.03, 95% CI, 3.94–101.82)), followed by BCG given post infancy (RR 8.13, 95% CI, 2.54–26.03)) and then to a lesser degree when BCG was given in infancy (RR 6.43, 95% CI, 1.72–24.85).

Interpretation

IGRA is feasible in Iqaluit, Nunavut, a remote Arctic community. IGRA testing results in more test results available to patients compared to TST. This test could result in fewer patients requiring latent TB treatment among those previously vaccinated with BCG in a region with limited public health human resources.     

Childhood Tuberculosis in Northern Viet Nam: A Review of 103 Cases

Background

Childhood tuberculosis causes significant morbidity and mortality in Southeast Asia, yet little is known about the epidemiology and clinical characteristics of this disease in Viet Nam.

Objectives

To determine the demographics, clinical presentations, radiographic and microbiologic findings, treatment regimens, and outcomes of children admitted with tuberculosis (TB) to a national referral hospital in Viet Nam.

Methods

We conducted a retrospective case series study of children ≤ 15 years old with bacteriologically confirmed or clinically diagnosed TB admitted to a national referral hospital in Ha Noi, Viet Nam from January through December 2007.

Results

One hundred three children were identified: median age 5 years (IQR 2-10), 44% female, 99% Kinh ethnicity, 27% residing in Ha Noi, 88% with BCG vaccination, 27% with known TB contact, and 38% malnourished. Intrathoracic TB was present in 62%, extrathoracic in 52%, both intra and extrathoracic in 19%, and undetermined site in 5%. The most common extrathoracic manifestation was peripheral lymphadenitis, and children under 5 were more likely to have miliary TB or both intra and extrathoracic TB. Fever and failure to thrive were common presenting symptoms among all participants (65% and 56%, respectively), 66% of those with intrathoracic TB presented with cough, and 92% of those with TB meningitis presented with severe neurologic impairment. Acid-fast bacilli smears and mycobacterial cultures were positive in 18% and 21% of children tested, and histopathology was positive in 88% of those biopsied. There were no adverse drug reactions necessitating change in therapy, and no inpatient mortality.

Conclusions

Extrathoracic TB was common, treatment well tolerated and clinical outcomes excellent. Culture confirmation rates were low and emphasize the need for improved diagnostics.