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Blood stream infections rank among the top seven causes of death of the general population. The aim of our study was to better understand the epidemiology of BSI in order to improve diagnostics and patient outcome. We used retrospective aggregated laboratory data of blood samples received from all public hospitals in Tyrol, Austria between 2006 and 2015. Microorganisms were categorized into obligatory, facultative, unusual pathogens and contaminants. The distribution, the cumulative incidence and antimicrobial susceptibility patterns were compared between the tertiary (TH) and regional peripheral hospitals (PH). Among 256,364 blood samples, 76.1% were from the TH The incidence of obligatory pathogens was 1.7 fold, and up to 3 times higher for facultative, unusual pathogens and contaminants in the TH and increased mainly due to an increase of E.coli, which was the most common isolated pathogen (n = 2,869), followed by Staphylococcus aureus (n = 1,439), Enterococcus sp. (n = 953) and Klebsiella sp. (n = 816). The distribution of obligatory pathogens differed between the hospital settings: In the TH Enterococcus sp. accounted for 40.8% and E.coli for 70.4%, respectively, whereas in the PH for 25.4% (p<0.0001) and 57.8%, respectively (p<0.0001) Antibiotic resistance of Gram negative bacteria and Staphylococcus aureus did not change during the observation period. Carbapenem resistance of Klebsiella sp. and vancomycin and linezolid resistance of Enterococcus faecium showed a non-significant increase since 2010 in the TH setting. We concluded that the incidence of BSI in TH was higher compared to PH. We observed higher contamination rates in the TH. We could not interpret the data of coagulase negative staphylococci due to lack of clinical data. We strongly recommend enhancement of training on blood culture sampling to decrease the rate of contamination. Due to differences in pathogen distribution and antimicrobial resistance between different hospital settings we recommend separate treatment guidelines for BSI by hospital setting. [ABSTRACT FROM AUTHOR]

Background: Linezolid has shown strong antimicrobial activity against multidrug-resistant (MDR)/rifampin-resistant strains of Mycobacterium tuberculosis. Linezolid achieves clinical efficacy mainly through area under the concentration time curve/minimum inhibitory concentration ratio in the infected lesion site. Previous studies mainly focused on the relationship between linezolid concentrations in the blood and infected bone tissue when the blood drug concentration reached the peak 2 h after administration. However, we do not know whether linezolid can maintain the same bone/plasma ratio in infected bone tissue when the blood concentration reaches the trough level. Therefore, this study aimed to evaluate the penetrability of linezolid into bone tissue 24 h after administration in patients with MDR spinal tuberculosis (TB). Methods: Nine MDR spinal TB patients, who received a treatment regimen including linezolid and underwent surgery, were enrolled prospectively from April 2017 to March 2019. Blood and diseased bone tissue specimens were collected simultaneously during operations 24 h after taking 600 mg of linezolid orally. Linezolid concentrations in plasma and diseased bone tissue specimens were determined by high-performance liquid chromatography–tandem mass spectrometry. Results: Following a 600 mg oral administration of linezolid 24 h before surgery, median concentrations of linezolid in plasma and diseased bone tissue for the 9 patients were 1.98 mg/L (range 0.30–3.44 mg/L) and 0.60 mg/L (range 0.18–2.13 mg/L), respectively, at resection time. The median diseased bone/plasma linezolid concentration ratio was 0.48 (range 0.30–0.67). Pearson’s correlation analysis showed that linezolid concentrations in the plasma were positively related to those in diseased bone tissue (r = 0.949, p < 0.001). Conclusions: After 24 h of medication, linezolid still had good penetrability into diseased bone tissue in patients with MDR spinal TB. [ABSTRACT FROM AUTHOR]

Background: Determine TB-LAM Ag (LAM) is a point of care test developed to diagnose tuberculosis (TB). The aim of this study was to evaluate the diagnostic performance of LAM in people living with HIV using Brazilian public health network algorithm for TB diagnosis. Methods and findings: A cross-sectional study design was used to enroll 199 adult patients in two sites in Rio de Janeiro and two in São Paulo. The study enrolled HIV-infected patients with CD4 counts ≤200 cells/mm3 (in the Alere PIMA CD4 assay at study screening), patients coughing for at least 2 weeks or presenting a chest radiography suggestive of TB. LAM, in conjunction with sputum smear microscopy or Xpert MTB/RIF (Xpert) as compared to Mycobacterium tuberculosis culture, which was used as a reference standard. TB prevalence was 24.6%. Overall accuracy of LAM was 79.9% (73.8%-84.9%), positive and negative predictive values were 62.2% (46.1%-75.9%) and 84% (77.5%-88.8%), respectively. The overall LAM sensitivity was 46.9% (33.7%-60.6%) and specificity was 90.7% (84.9%-94.4%). The best performance of LAM was observed among patients with CD4 counts ≤50 cells/mm3 (sensitivity = 70.4% and specificity = 85.9%). When 2 respiratory smears were used in conjunction with LAM, sensitivity increased 22%, as compared to just 2 smears. Furthermore, LAM when used in conjunction with two respiratory smears, was as sensitive as compared to a single one. However, no improvement in TB diagnosis occurred when LAM was used with Xpert as compared to Xpert alone. Among 14 LAM false positive tests, Non-Tuberculosis Mycobacteria were isolated in three cases. Conclusion: LAM is a point of care test that increased TB diagnosis in immunosuppressed HIV-infected patients when used in conjunction with smear microscopy, but not when used with Xpert in Brazilian public health network sites. Use of LAM test should be considered in settings where immunosuppressed HIV patients need rapid TB diagnosis. [ABSTRACT FROM AUTHOR]

The survey described in this research paper aimed to investigate the economic and health impacts of birds on dairies. Birds are common pests on dairies, consuming and contaminating feed intended for cattle. As a result, dairy operators experience increased feed costs and increased pathogen and disease risk. We surveyed dairy operators attending the 2017 Washington Dairy Conference to examine the impact of birds on dairies in Washington State. Dairy operators reported feed losses valued at $55 per cow resulting in annual losses totaling $5.5 million in the Western region of the state and $9.2 million in the Eastern region of the state. Shooting was the most commonly used bird management method and European starlings (Sternus vulgaris) were the most frequently implicated species statewide. Bird abundance greater than 10,000 birds per day was associated with larger herd size and with self-reported presence of Johne’s disease and Salmonella. [ABSTRACT FROM AUTHOR]

Introduction: In Tunisia, almost 77% of clinically and bacteriologically diagnosed cases of extrapulmonary tuberculosis (EPTB) are zoonotic TB, caused by M. bovis. Although several studies have analyzed bovine TB in cattle in Tunisia, no study has evaluated the risk of transmission to humans in such an endemic country. We aimed to study the genetic diversity of M. bovis human isolates, to ascertain the causes of human EPTB infection by M. bovis and to investigate the distribution and population structure of this species in Tunisia. Materials and methods: A total of 110 M. bovis isolates taken from patients with confirmed EPTB were characterized by spoligotyping and MIRU-VNTR typing methods. Results: Among the 15 spoligotypes detected in our study, 6 (SB0120, SB0121, SB2025, SB1200, SB1003 and SB0134) were the most prevalent (83.5%) of which SB0120, SB0121 and SB2025 were the most prevailing. MIRU-VNTR typing method showed a high genotypic and genetic diversity. The genetic differentiation based on MIRU-VNTR was significant between populations from South East (Tataouine, Medenine) and Central West (Gafsa, Sidi Bouzid, Kasserine) regions. Of note, 13/15 (86.7%) spoligotypes detected in our study were previously identified in cattle in Tunisia with different frequencies suggesting a peculiar ability of some genotypes to infect humans. Using combined spoligotyping and MIRU-VNTR method, a high clustering rate of 43.9% was obtained. Our results underlined that human EPTB due to M. bovis was more commonly found in female gender and in young patients. Most of our patients, 66.4% (73/110) were raw milk or derivatives consumers, whereas 30.9% (34/110) patients would have contracted EPTB through contact with livestock. The findings suggest that the transmission of Zoonotic TB caused by M. bovis to humans mainly occurred by oral route through raw milk or derivatives. Conclusion: Our study showed the urgent need of a better veterinary control with the implementation of effective and comprehensive strategies in order to reach a good protection of animals as well as human health. [ABSTRACT FROM AUTHOR]

Sexually transmitted infections (STIs) remain a worldwide problem and a severe threat to public health. The purpose of this study was to compare Aptima® Assays (Hologic®) and the Allplex™ STI Essential Assay (Seegene®) for the simultaneous detection of Chlamydia trachomatis, Neisseria gonorrhoeae, Trichomonas vaginalis and Mycoplasma genitalium in clinical practice. The Aptima® assays (Hologic®) are based on a transcription-mediated amplification (TMA) method. The Allplex™ STI Essential assay (Seegene®) is based on a multiplex Real-Time PCR (RT-PCR) method. A total of 622 clinical samples from different anatomical sites were tested using both methods. A total of 88 (14.1%) and 66 (10.6%) positive samples were found for any of the TMA assays used and for the RT-PCR assay, respectively. Aptima® assays showed a slightly higher rate of positive results for all pathogens except for T. vaginalis, the results of which were similar to those obtained with Allplex™. The most commonly detected pathogen was C. trachomatis (37 samples; 5.9% using TMA assays) and the anatomical site with the highest prevalence of microorganisms was a non-urogenital site, the pharynx (27 positive samples; 4.3%). Using the Aptima® assays as reference method, the comparison showed that the average specificity of multiplex RT-PCR was 100.0% for the four pathogens. However an average sensitivity of 74.5% was observed, showing 95.2% (CI95%; 93.6–96.9) of overall concordance (κ = 0.80). In conclusion, the Aptima® assays show a higher sensitivity on a wide range of sample types compared to the Allplex™ assay. [ABSTRACT FROM AUTHOR]

Background: Limited data existed exclusively describing Mycobacterium tuberculosis lineage 3 (MTB-L3), sub-lineages, and clinical manifestations in Kampala, Uganda. This study sought to elucidate the circulating MTB-L3 sub-lineages and their corresponding clinical phenotypes. Method: A total of 141 M. tuberculosis isolates were identified as M. tuberculosis lineage 3 using Single nucleotide polymorphism (SNP) marker analysis method. To ascertain the sub-lineages/sub-strains within the M. tuberculosis lineage 3, the direct repeat (DR) loci for all the isolates was examined for sub-lineage specific signatures as described in the SITVIT2 database. The infecting sub-strains were matched with patients’ clinical and demographic characteristics to identify any possible association. Result: The data showed 3 sub-lineages circulating with CAS 1 Delhi accounting for 55% (77/141), followed by CAS 1-Kili 16% (22/141) and CAS 2/CAS 8% (12/141). Remaining isolates 21% (30/141) were unclassifiable. To explore whether the sub-lineages differ in their ability to cause increased severe disease, we used extent of lung involvement as a proxy for severe disease. Multivariable analysis showed no association between M. tuberculosis lineage 3 sub-lineages with severe disease. The risk factors associated with severe disease include having a positive smear (OR = 9.384; CI 95% = 2.603–33.835), HIV (OR = 0.316; CI 95% = 0.114–0.876), lymphadenitis (OR = 0. 171; CI 95% = 0.034–0.856) and a BCG scar (OR = 0.295; CI 95% = 0.102–0.854). Conclusion: In Kampala, Uganda, there are three sub-lineages of M. tuberculosis lineage 3 that cause disease of comparable severity with CAS-Dehli as the most prevalent. Having HIV, lymphadenitis, a BCG scar and a smear negative status is associated with reduced severe disease. [ABSTRACT FROM AUTHOR]

Nymphaea tetragona Georgi (Nymphaceae) is traditionally used in Asia for the treatment of diarrhea, dysentery and fever. The plant contains various active compounds, including methyl gallate (MG) which are reported to inhibit bacterial virulence mechanisms. This study aimed to evaluate the alterations on viability, membrane potential and integrity of Salmonella enterica Serovar Typhimurium exposed to MG in combination with Tylosin (Ty), which is relatively inactive against Gram-negative bacteria, but it is commonly used as a feed additive in livestock. Besides, the effects of sub-inhibitory concentrations of the combination (MT) on the interaction between S. Typhimurium and the host cell, as well as on the indirect host responses, were characterized. Flow cytometry, confocal and electron microscopic examinations were undertaken to determine the effects of MT on S. Typhimurium. The impacts of sub-inhibitory concentrations of MT on biofilm formation, as well as on the adhesion, invasion and intracellular survival of S. Typhimurium were assessed. The result demonstrated significant damage to the bacterial membrane, leakage of cell contents and a reduction in the membrane potential when treated with MT. Sub-inhibitory concentrations of MT significantly reduced (P < 0.05) the biofilm-forming, adhesive and invasive abilities of S. Typhimurium. Exposure to MT drastically reduced the bacterial count in macrophages. Up-regulation of interleukin (IL)-6, IL-8 and IL-10 cytokine genes were detected in intestinal epithelial cells pre-treated with MT. This report is the first to describe the effects of MT against S. Typhimurium. The result indicates a synergistic interaction between MG and Ty against S. Typhimurium. Therefore, the combination may be a promising option to combat S. Typhimurium in swine and, indirectly, safeguard the health of the public. [ABSTRACT FROM AUTHOR]

The anticariogenic activity of the extract of Glycyrrhiza uralensis (licorice) has been well documented. We recently developed an herbal lollipop containing licorice extracts with Glycyrrhizol A, the compound displaying strong antimicrobial activity against Streptococcus mutans. Preliminary testing showed that the herbal lollipop reduced salivary S. mutans counts in vivo. In this study, we aimed to further test the efficacy of this herbal lollipop for reducing salivary S. mutans levels, and investigate its impact on salivary microbiome. Using a well-established in vitro oral microbiome model, we showed that licorice extract displays targeted killing against S. mutans without affecting the biodiversity of the community. In vivo study corroborated in vitro findings, showing for high caries-risk children aged 3–6 with salivary S. mutans levels >5x105 cells/ml, daily use of 2 licorice-containing lollipops for 3 weeks significantly reduced salivary S. mutans levels compared to the control group. Salivary microbiome analysis showed either no change or even increase in phylogenetic diversity of the oral community following herbal lollipop usage. Although further study with longer term observation is needed, these results suggest that use of licorice extract-containing lollipops can be as a simple and effective way to reduce the risk of dental caries in children. [ABSTRACT FROM AUTHOR]

Enteric fevers, caused by the Salmonella enterica serovars Typhi (ST), Paratyphi A (PA) and Paratyphi B (PB), are life-threatening illnesses exhibiting very similar clinical symptoms but with distinct epidemiologies, geographical distributions and susceptibilities to antimicrobial treatment. Nevertheless, the mechanisms by which the host recognizes pathogens with high levels of homology, such as these bacterial serovars, remain poorly understood. Using a three-dimensional organotypic model of the human intestinal mucosa and PA, PB, and ST, we observed significant differences in the secretion patterns of pro-inflammatory cytokines and chemokines elicited by these serovars. These cytokines/chemokines were likely to be co-regulated and influenced the function of epithelial cells, such as the production of IL-8. We also found differing levels of polymorphonuclear leukocyte (PMN) migration among various infection conditions that either included or excluded lymphocytes and macrophages (Mϕ), strongly suggesting feedback mechanisms among these cells. Blocking experiments showed that IL-1β, IL-6, IL-8, TNF-α and CCL3 cytokines were involved in the differential regulation of migration patterns. We conclude that the crosstalk among the lymphocytes, Mϕ, PMN and epithelial cells is cytokine/chemokine-dependent and bacterial-serotype specific, and plays a pivotal role in orchestrating the functional efficiency of the innate cells and migratory characteristics of the leukocytes. [ABSTRACT FROM AUTHOR]

This study investigated the prevalence of Orientia tsutsugamushi, Anaplasma phagocytophilum, and Leptospira interrogans in wild rodents through molecular detection using organ samples and through serological assay using blood samples of mice collected from two distinct sites in Gwangju Metropolitan City, Republic of Korea (ROK). A total of 47 wild rodents, identified as Apodemus agrarius (A. agrarius), were captured from June to August 2016. The seroprevalence of antibodies against bacterial pathogens in A. agrarius sera was analyzed; 17.4% (8/46) were identified as O. tsutsugamushi through indirect immunofluorescence assay and 2.2% (1/46) were identified as Leptospira species through passive hemagglutination assay. Using polymerase chain reaction, the spleen, kidney and blood samples were investigated for the presence of O. tsutsugamushi, A. phagocytophilum, and L. interrogans. Out of the 47 A. agrarius, 19.1% (9/47) were positive for A. phagocytophilum and 6.4% (3/47) were positive for L. interrogans, while none were positive for O. tsutsugamushi. Four out of 46 (8.7%) blood samples, six out of 45 (13.3%) spleen samples, and one out of 47 (2.1%) kidney samples were positive for A. phagocytophilum. Three out of 47 (6.4%) kidney samples were positive for L. interrogans. The sequencing results of PCR positive samples demonstrated > 99% similarity with A. phagocytophilum and L. interrogans sequences. A. phagocytophilum was mostly detected in the spleen, whereas L. interrogans was mostly detected in the kidneys. Notably, A. phagocytophilum and L. interrogans were detected in A. agrarius living in close proximity to humans in the metropolitan suburban areas. The results of this study indicate that rodent-borne bacteria may be present in wild rodents in the metropolitan suburban areas of ROK. [ABSTRACT FROM AUTHOR]

India has the highest burden of Tuberculosis (TB) and multidrug-resistant TB (MDR-TB) worldwide. Innovative technology is the need of the hour to identify these cases that remain either undiagnosed or inadequately diagnosed due to the unavailability of appropriate tools at primary healthcare settings. We developed and evaluated 3 kits, namely ‘TB Detect’ (containing BioFM-Filter device), ‘TB Concentration and Transport’ (containing Trans-Filter device) and ‘TB DNA Extraction’ kits. These kits enable bio-safe equipment-free concentration of sputum on filters and improved fluorescence microscopy at primary healthcare centres, ambient temperature transport of dried inactivated sputum filters to central laboratories and molecular detection of drug resistance by PCR and DNA sequencing (Mol-DST). In a 2-site evaluation (n = 1190 sputum specimens) on presumptive TB patients, BioFM-Filter smear exhibited a significant increase in positivity of 7% and 4% over ZN smear and LED-FM smear (p<0.05), respectively and an increment in smear grade status (1+ or 2+ to 3+) of 16% over ZN smear and 20% over LED-FM smear. The sensitivity of Mol-DST in presumptive MDR-TB and XDR-TB cases (n = 148) was 90% for Rifampicin (95% confidence interval [CI], 78–96%), 84% for Isoniazid (95% CI, 72–92%), 83% for Fluoroquinolones (95% CI, 66–93%) and 75% for Aminoglycosides (95% CI, 35–97%), using phenotypic DST as the reference standard. Test specificity was 88–93% and concordance was ~89–92% (κ value 0.8–0.9). The patient-friendly kits described here address several of the existing challenges and are designed to provide ‘Universal Access’ to rapid TB diagnosis, including drug-resistant disease. Their utility was demonstrated by application to sputum at 2 sites in India. Our findings pave the way for larger studies in different point-of-care settings, including high-density urban areas and remote geographical locations. [ABSTRACT FROM AUTHOR]

Streptococcus pneumoniae (pneumococci) is a leading cause of severe bacterial meningitis in many countries worldwide. To characterize the repertoire of fitness and virulence factors predominantly expressed during meningitis we performed niche-specific analysis of the in vivo proteome in a mouse meningitis model, in which bacteria are directly inoculated into the cerebrospinal fluid (CSF) cisterna magna. We generated a comprehensive mass spectrometry (MS) spectra library enabling bacterial proteome analysis even in the presence of eukaryotic proteins. We recovered 200,000 pneumococci from CSF obtained from meningitis mice and by MS we identified 685 pneumococci proteins in samples from in vitro filter controls and 249 in CSF isolates. Strikingly, the regulatory two-component system ComDE and substrate-binding protein AliB of the oligopeptide transporter system were exclusively detected in pneumococci recovered from the CSF. In the mouse meningitis model, AliB-, ComDE-, or AliB-ComDE-deficiency resulted in attenuated meningeal inflammation and disease severity when compared to wild-type pneumococci indicating the crucial role of ComDE and AliB in pneumococcal meningitis. In conclusion, we show here mechanisms of pneumococcal adaptation to a defined host compartment by a proteome-based approach. Further, this study provides the basis of a promising strategy for the identification of protein antigens critical for invasive disease caused by pneumococci and other meningeal pathogens. [ABSTRACT FROM AUTHOR]

Background: Clinical tuberculosis diagnosis and empiric treatment have traditionally been common among patients with negative bacteriologic test results. Increasing availability of rapid molecular diagnostic tests, including Xpert MTB/RIF and the new Xpert Ultra cartridge, may alter the role of empiric treatment. Methods: We prospectively enrolled outpatients age > = 15 who were evaluated for pulmonary tuberculosis at three health facilities in Kampala, Uganda. Using sputum mycobacterial culture, interviews, and clinical record abstraction, we estimated the accuracy of clinical diagnosis relative to Xpert and sputum culture and assessed the contribution of clinical diagnosis to case detection. Results: Over a period of 9 months, 99 patients were diagnosed with pulmonary tuberculosis and subsequently completed sputum culture; they were matched to 196 patients receiving negative tuberculosis evaluations in the same facilities. Xpert was included in the evaluation of 291 (99%) patients. Compared to culture, Xpert had a sensitivity of 92% (95% confidence interval 83–97%) and specificity of 95% (92–98%). Twenty patients with negative Xpert were clinically diagnosed with tuberculosis and subsequently had their culture status determined; two (10%) were culture-positive. Considering all treated patients regardless of Xpert and culture data completeness, and considering treatment initiations before a positive Xpert (N = 4) to be empiric, 26/101 (26%) tuberculosis treatment courses were started empirically. Compared to sputum smear- or Xpert-positive patients with positive cultures, empirically-treated, Xpert-negative patients with negative cultures had higher prevalence of HIV (67% versus 37%), shorter duration of cough (median 4 versus 8 weeks), and lower inflammatory markers (median CRP 7 versus 101 mg/L). Conclusion: Judged against sputum culture in a routine care setting of high HIV prevalence, the accuracy of Xpert was high. Clinical judgment identified a small number of additional culture-positive cases, but with poor specificity. Although clinicians should continue to prescribe tuberculosis treatment for Xpert-negative patients whose clinical presentations strongly suggest pulmonary tuberculosis, they should also carefully consider alternative diagnoses. [ABSTRACT FROM AUTHOR]

Typhoid fever is endemic across sub-Saharan Africa. However, estimates of the burden of typhoid are undermined by insufficient blood volumes and lack of sensitivity of blood culture. Here, we aimed to address this limitation by exploiting pre-enrichment culture followed by PCR, alongside routine blood culture to improve typhoid case detection. We carried out a prospective diagnostic cohort study and enrolled children (aged 0–4 years) with non-specific febrile disease admitted to a tertiary hospital in Blantyre, Malawi from August 2014 to July 2016. Blood was collected for culture (BC) and real-time PCR after a pre-enrichment culture in tryptone soy broth and ox-bile. DNA was subjected to PCR for invA (Pan-Salmonella), staG (S. Typhi), and fliC (S. Typhimurium) genes. A positive PCR was defined as invA plus either staG or fliC (CT<29). IgM and IgG ELISA against four S. Typhi antigens was also performed. In total, 643 children (median age 1.3 years) with nonspecific febrile disease were enrolled; 31 (4.8%) were BC positive for Salmonella (n = 13 S. Typhi, n = 16 S. Typhimurium, and n = 2 S. Enteritidis). Pre-enrichment culture of blood followed by PCR identified a further 8 S. Typhi and 15 S. Typhimurium positive children. IgM and IgG titres to the S. Typhi antigen STY1498 (haemolysin) were significantly higher in children that were PCR positive but blood culture negative compared to febrile children with all other non-typhoid illnesses. The addition of pre-enrichment culture and PCR increased the case ascertainment of invasive Salmonella disease in children by 62–94%. These data support recent burden estimates that highlight the insensitivity of blood cultures and support the targeting of pre-school children for typhoid vaccine prevention in Africa. Blood culture with real-time PCR following pre-enrichment should be used to further refine estimates of vaccine effectiveness in typhoid vaccine trials. [ABSTRACT FROM AUTHOR]

Background: Current diagnostics are inadequate to meet the challenges presented by co-infection with Mycobacterium tuberculosis (Mtb) and HIV, the leading cause of death for HIV-infected individuals. Improved characterization of Mtb/HIV coinfection as a distinct disease state may lead to better identification and treatment of affected individuals. Methods: Four previously-published TB and HIV co-infection related datasets were used to train and validate multinomial machine learning classifiers that simultaneously predict TB and HIV status. Classifier predictive performance was measured using leave-one-out cross validation on the training set and blind predictive performance on multiple test sets using area under the ROC curve (AUC) as the performance metric. Linear modelling of signature gene expression was applied to systematically classify genes as TB-only, HIV-only or combined TB/HIV. Results: The optimal signature discovered was a 10-gene random forest multinomial signature that robustly discriminated active tuberculosis (TB) from other non-TB disease states with improved performance compared with previously published signatures (AUC: 0.87), and specifically discriminated active TB/HIV co-infection from all other conditions (AUC: 0.88). Signature genes exhibited a variety of transcriptional patterns including both TB-only and HIV-only response genes and genes with expression patterns driven by interactions between HIV and TB infection states, including the CD8+ T-cell receptor LAG3 and the apoptosis-related gene CERKL. Conclusions: By explicitly including distinct disease states within the machine learning analysis framework, we developed a compact and highly diagnostic signature that simultaneously discriminates multiple disease states associated with Mtb/HIV co-infection. Examination of the expression patterns of signature genes suggests mechanisms underlying the unique inflammatory conditions associated with active TB in the presence of HIV. In particular, we observed that dysregulation of CD8+ effector T-cell and NK-cell associated genes may be an important feature of Mtb/HIV co-infection. [ABSTRACT FROM AUTHOR]

Persistent bacteremia caused by Staphylococcus aureus (SA), especially methicillin-resistant SA (MRSA), is a significant cause of morbidity and mortality. Despite susceptibility phenotypes in vitro, persistent MRSA strains fail to clear with appropriate anti-MRSA therapy during bacteremia in vivo. Thus, identifying the factors that cause such MRSA persistence is of direct and urgent clinical relevance. To address the dynamics of MRSA persistence in the face of host immunity and typical antibiotic regimens, we developed a mathematical model based on the overarching assumption that phenotypic heterogeneity is a hallmark of MRSA persistence. First, we applied an ensemble modeling approach and obtained parameter sets that satisfied the condition of a minimum inoculum dose to establish infection. Second, by simulating with the selected parameter sets under vancomycin therapy which follows clinical practices, we distinguished the models resulting in resolving or persistent bacteremia, based on the total SA exceeding a detection limit after five days of treatment. Third, to find key determinants that discriminate resolving and persistent bacteremia, we applied a machine learning approach and found that the immune clearance rate of persister cells is a key feature. But, fourth, when relapsing bacteremia was considered, the growth rate of persister cells was also found to be a key feature. Finally, we explored pharmacological strategies for persistent and relapsing bacteremia and found that a persister killer, but not a persister formation inhibitor, could provide for an effective cure the persistent bacteremia. Thus, to develop better clinical solutions for MRSA persistence and relapse, our modeling results indicate that we need to better understand the pathogen-host interactions of persister MRSAs in vivo. [ABSTRACT FROM AUTHOR]

Melioidosis is a severe infectious disease caused by gram-negative, facultative intracellular pathogen Burkholderia pseudomallei (B. pseudomallei). Although cases are increasing reported from other parts of the world, it is an illness of tropical and subtropical climates primarily found in southeast Asia and northern Australia. Because of a 40% mortality rate, this life-threatening disease poses a public health risk in endemic area. Early detection of B. pseudomallei infection is vital for prognosis of a melioidosis patient. In this study, a novel isothermal recombinase polymerase amplification combined with lateral flow dipstick (LF-RPA) assay was established for rapid detection of B. pseudomallei. A set of primer-probe targeting orf2 gene within the putative type III secretion system (T3SS) cluster genes was generated and parameters for the LF-RPA assay were optimized. Result can be easy visualized in 30 minutes with the limit of detection (LOD) as low as 20 femtogram (fg) (ca. 25.6 copies) of B. pseudomallei genomic DNA without a specific equipment. The assay is highly specific as no cross amplification was observed with Burkholderia mallei, members of the Burkholderia cepacia-complex and 35 non-B. pseudomallei bacteria species. Moreover, isolates from patients in Hainan (N = 19), Guangdong (N = 1), Guangxi (N = 3) province of China as well as in Australia (N = 3) and Thailand (N = 1) were retrospectively confirmed by the newly developed method. LODs for B. pseudomallei-spiked soil and blood samples were 2.1×103 CFU/g and 4.2×103 CFU/ml respectively. The sensitivity of the LF-RPA assay was comparable to TaqMan Real-Time PCR (TaqMan PCR). In addition, the LF-RPA assay exhibited a better tolerance to inhibitors in blood than TaqMan PCR. Our results showed that the LF-RPA assay is an alternative to existing PCR-based methods for detection of B. pseudomallei with a potentiality of early accurate diagnosis of melioidosis at point of care or in-field use. [ABSTRACT FROM AUTHOR]

Background: In high-burden settings, case fatality rates are reported to be between 11% and 32% in hospitalized patients with HIV-associated tuberculosis, yet the underlying causes of mortality remain poorly characterized. Understanding causes of mortality could inform the development of novel management strategies to improve survival. We aimed to assess clinical and microbiologic determinants of mortality and to characterize the pathophysiological processes underlying death by evaluating host soluble inflammatory mediators and determined the relationship between these mediators and death as well as biomarkers of disseminated tuberculosis.Methods and Findings: Adult patients with HIV hospitalized with a new diagnosis of HIV-associated tuberculosis were enrolled in Cape Town between 2014 and 2016. Detailed tuberculosis diagnostic testing was performed. Biomarkers of tuberculosis dissemination and host soluble inflammatory mediators at baseline were assessed. Of 682 enrolled participants, 576 with tuberculosis (487/576, 84.5% microbiologically confirmed) were included in analyses. The median age was 37 years (IQR = 31-43), 51.2% were female, and the patients had advanced HIV with a median cluster of differentiation 4 (CD4) count of 58 cells/L (IQR = 21-120) and a median HIV viral load of 5.1 log10 copies/mL (IQR = 3.3-5.7). Antituberculosis therapy was initiated in 566/576 (98.3%) and 487/576 (84.5%) started therapy within 48 hours of enrolment. Twelve-week mortality was 124/576 (21.5%), with 46/124 (37.1%) deaths occurring within 7 days of enrolment. Clinical and microbiologic determinants of mortality included disseminated tuberculosis (positive urine lipoarabinomannan [LAM], urine Xpert MTB/RIF, or tuberculosis blood culture in 79.6% of deaths versus 60.7% of survivors, p = 0.001), sepsis syndrome (high lactate in 50.8% of deaths versus 28.9% of survivors, p < 0.001), and rifampicin-resistant tuberculosis (16.9% of deaths versus 7.2% of survivors, p = 0.002). Using non-supervised two-way hierarchical cluster and principal components analyses, we describe an immune profile dominated by mediators of the innate immune system and chemotactic signaling (interleukin-1 receptor antagonist [IL-1Ra], IL-6, IL-8, macrophage inflammatory protein-1 beta [MIP-1β]/C-C motif chemokine ligand 4 [CCL4], interferon gamma-induced protein-10 [IP-10]/C-X-C motif chemokine ligand 10 [CXCL10], MIP-1 alpha [MIP-1α]/CCL3), which segregated participants who died from those who survived. This immune profile was associated with mortality in a Cox proportional hazards model (adjusted hazard ratio [aHR] = 2.2, 95%CI = 1.9-2.7, p < 0.001) and with detection of biomarkers of disseminated tuberculosis. Clinicians attributing causes of death identified tuberculosis as a cause or one of the major causes of death in 89.5% of cases. We did not perform longitudinal sampling and did not have autopsy-confirmed causes of death.Conclusions: In this study, we did not identify a major contribution from coinfections to these deaths. Disseminated tuberculosis, sepsis syndrome, and rifampicin resistance were associated with mortality. An immune profile dominated by mediators of the innate immune system and chemotactic signaling was associated with both tuberculosis dissemination and mortality. These findings provide pathophysiologic insights into underlying causes of mortality and could be used to inform the development of novel treatment strategies and to develop methods to risk stratify patients to appropriately target novel interventions. Causal relationships cannot be established from this study. [ABSTRACT FROM AUTHOR]

Despite reducing the prevalent foodborne pathogen Campylobacter jejuni in chickens decreases campylobacteriosis, few effective approaches are available. The aim of this study was to use microbial metabolic product bile acids to reduce C. jejuni chicken colonization. Broiler chicks were fed with deoxycholic acid (DCA), lithocholic acid (LCA), or ursodeoxycholic acid (UDCA). The birds were also transplanted with DCA modulated anaerobes (DCA-Anaero) or aerobes (DCA-Aero). The birds were infected with human clinical isolate C. jejuni 81–176 or chicken isolate C. jejuni AR101. Notably, C. jejuni 81–176 was readily colonized intestinal tract at d16 and reached an almost plateau at d21. Remarkably, DCA excluded C. jejuni cecal colonization below the limit of detection at 16 and 28 days of age. Neither chicken ages of infection nor LCA or UDCA altered C. jejuni AR101 chicken colonization level, while DCA reduced 91% of the bacterium in chickens at d28. Notably, DCA diet reduced phylum Firmicutes but increased Bacteroidetes compared to infected control birds. Importantly, DCA-Anaero attenuated 93% of C. jejuni colonization at d28 compared to control infected birds. In conclusion, DCA shapes microbiota composition against C. jejuni colonization in chickens, suggesting a bidirectional interaction between microbiota and microbial metabolites. [ABSTRACT FROM AUTHOR]

The prevalence of rheumatic heart disease (RHD) in the Aboriginal population of the Australian Northern Territory is high, and Streptococcus pyogenes skin infections likely contribute to this. A promising candidate S. pyogenes “30mer” vaccine is composed of 30 pharyngitis associated type-specific antigens from the S. pyogenes M protein. Cross opsonisation experiments suggest that 30mer vaccine protection may extend to non-cognate emm types. A new “emm cluster” scheme for classifying M protein is based on the full-length coding sequence, and correlates with functional and immunological properties, and anatomical tropism. Twenty-seven years of research in the Northern Territory has yielded 1810 S. pyogenes isolates with clinical and emm type data. The primary aim was to analyse these data with reference to the emm cluster scheme and cross opsonisation information, to inform estimation of 30mer vaccine efficacy in the Northern Territory. The isolates encompass 101 emm types. Variants of cluster A-C were enriched in throat isolates, and variants of emm cluster D enriched in skin isolates. Throat isolates were enriched for 30mer vaccine cognate emm types in comparison with skin isolates of which only 25% were vaccine emm types. While cross opsonisation data indicates potential for enhancing 30mer vaccine coverage, more than one third of skin isolates were within 38 emm types untested for cross opsonisation. Emm cluster D variants, in particular emm cluster D4, were not only all non-cognate with the vaccine, but were abundant and diverse, and less likely to be cross-opsonisation positive than other emm clusters. Long term persistence of many emm types in the study area was revealed. It was concluded that the 30mer vaccine efficacy in the Northern Territory will likely require both cross protection, and additional measures to elicit immunity against variants of emm cluster D. [ABSTRACT FROM AUTHOR]

Background: From 2014 to 2017, the number of migrants who came to Italy via the Mediterranean route has reached an unprecedented level. The majority of refugees and migrants were rescued in the Central Mediterranean and disembarked at ports in the Sicily region. Rapid on-spot active TB screening intervention at the point of arrival will cover most migrants arriving in EU and by detecting TB prevalent cases will limit further transmission of the disease. Material and methods: Between November 2016 and December 2017 newly arrived migrants at point of arrivals in Sicily, were screened for active Tuberculosis using a smartphone application, followed in symptomatic individuals by fast molecular test, Xpert MTB/RIF Ultra, on collected sputum samples. Results: In the study period 3787 migrants received a medical evaluation. Eight hundred and ninety-one (23.5%) reported at least one protocol-defined Tuberculosis symptom. Fifteen (2.7%) were positive to at least one microbiological test revealing a post-entry screening prevalence rate of 396 per 100.000 individuals screened (95% CI: 2.22–6.53). In logistic regression analysis, those with cough and at least one other symptom had an increased probability of testing positive compared to persons with symptoms other than cough. Whole-genome-sequencing demonstrate two separate cases of transmission. Discussion: To our knowledge this study reports first-time results of an active TB case finding strategy based on on-spot symptom screening using a smartphone application, followed by fast molecular test on collected sputum samples. Our preliminary findings reveal a post-entry screening prevalence rate of 396 per 100.000 individuals screened (95% CI: 2.22–6.53). [ABSTRACT FROM AUTHOR]

The current study measured the influence of milk of subclinically infected glands by different bacteria species on the cow’s milk. The effects of bacterial infection or inflammation on gland milk yield were related to the bacteria species that caused the infection. The volume of milk of the inflamed gland from the cow’s milk yield was significantly lower (P<0.001) for the glands previously infected by Escherichia coli (PIEc) and those infected with Streptococcus dysgalactiae. Coagulation properties, rennet clotting time (RCT) and curd firmness (CF) also depended on the bacteria causing the infection. RCT values of all the inflamed glands were significantly longer (P<0.001) and CF values were significantly lower than that of the healthy ones. Moreover, in the whole milk, CF was also significantly lower and not proportional to the volume of the milk from the inflamed gland of the cow’s milk. Calculation of the predicted 40% dry matter curd weight (PCW) on the cow level, including the healthy and inflamed glands or the healthy glands alone, showed that for 10 of 13 PIEc cows, the presence of the affected gland’s milk in the whole cow milk resulted in a lower PCW value. Likewise, 7 of 20 cows infected by S. dysgalactiae had negative delta values. Unlike the latter bacteria, PCW from milk of glands infected with CNS increased, although in a lower magnitude than in the healthy glands. No correlation was found between logSCC in the whole cow milk (healthy and inflamed glands) and PCW. [ABSTRACT FROM AUTHOR]

Background: Simple methods for the accurate triaging and screening of HIV-associated tuberculosis (TB) are urgently needed. We hypothesized that combining serum antibody with urine lipoarabinomannan (U-LAM) detection can improve the detection of HIV-associated TB. Methods: We performed a case-control study with sampling from a prospective study of South African HIV-infected subjects who were screened for TB prior to initiating antiretroviral therapy. Sera from all available TB cases (n = 74) and randomly selected non-TB controls (n = 30), all tested for U-LAM, sputum microscopy, GeneXpert, and cultures, were evaluated for antibodies to LAM and arabinomannan (AM). Diagnostic logistic regression models for TB were developed based on the primary test results and the additive effect of antibodies with leave-one-out cross-validation. Results: Antibody responses to LAM and AM correlated strongly (p<0.0001), and IgG and IgM reactivities were significantly higher in TB than non-TB patients (p<0.0001). At 80% specificity, the target specificity for a non-sputum-based simple triage/screening test determined by major TB stakeholders, combining U-LAM with IgG detection significantly increased the sensitivity for HIV-associated TB to 92% compared to 30% for U-LAM alone (p<0.001). Sputum microscopy combined with IgG detection increased sensitivity to 88% compared to 31% for microscopy alone, and Xpert with IgG increased sensitivity to 96% and 99% compared to 57% for testing one, and 70% for testing two sputa with Xpert alone, respectively. Conclusion: Combining U-LAM with serum antibody detection could provide a simple low-cost method that meets the requirements for a non-sputum-based test for the screening of HIV-associated TB. [ABSTRACT FROM AUTHOR]

Background: Salmonella enterica serovar Enteritidis is a cause of both poultry- and egg-associated enterocolitis globally and bloodstream-invasive nontyphoidal Salmonella (iNTS) disease in sub-Saharan Africa (sSA). Distinct, multi-drug resistant genotypes associated with iNTS disease in sSA have recently been described, often requiring treatment with fluoroquinolone antibiotics. In industrialised countries, antimicrobial use in poultry production has led to frequent fluoroquinolone resistance amongst globally prevalent enterocolitis-associated lineages. Methodology/Principal findings: Twenty seven S. Enteritidis isolates from patients with iNTS disease and two poultry isolates, collected between 2007 and 2015 in the Ashanti region of Ghana, were whole-genome sequenced. These isolates, notable for a high rate of diminished ciprofloxacin susceptibility (DCS), were placed in the phyletic context of 1,067 sequences from the Public Health England (PHE) S. Enteritidis genome database to understand whether DCS was associated with African or globally-circulating clades of S. Enteritidis. Analysis showed four of the major S. Enteritidis clades were represented, two global and two African. All thirteen DCS isolates, containing a single gyrA mutation at codon 87, belonged to a global PT4-like clade responsible for epidemics of poultry-associated enterocolitis. Apart from two DCS isolates, which clustered with PHE isolates associated with travel to Spain and Brazil, the remaining DCS isolates, including one poultry isolate, belonged to two monophyletic clusters in which gyrA 87 mutations appear to have developed within the region. Conclusions/Significance: Extensive phylogenetic diversity is evident amongst iNTS disease-associated S. Enteritidis in Ghana. Antimicrobial resistance profiles differed by clade, highlighting the challenges of devising empirical sepsis guidelines. The detection of fluoroquinolone resistance in phyletically-related poultry and human isolates is of major concern and surveillance and control measures within the region’s burgeoning poultry industry are required to protect a human population at high risk of iNTS disease. [ABSTRACT FROM AUTHOR]

Canine brucellosis, caused by Brucella canis, is a disease of dogs and represents a public health concern as it can be transmitted to humans. Canine brucellosis is on the rise in the United States and there is currently no vaccine for use in dogs. Mice have been extensively utilized to investigate host-pathogen interactions and vaccine candidates for smooth Brucella species and could serve a similar role for studying B. canis. However, comparatively little is known about B. canis infection in mice. The objective of this study was to characterize the kinetics of colonization and pathogenicity of B. canis in mice in order to evaluate the mouse as a model for studying this pathogen. C57BL/6 mice were inoculated intraperitoneally with 105, 107, or 109 CFU of Brucella canis RM6/66 and euthanized 1-, 2-, 4-, 6-, 9-, and 12-weeks post-inoculation. B. canis induced splenomegaly in mice infected with 109 CFU at 1- and 2 weeks post-inoculation while no gross lesions were observed in other dose groups. Infection at the two higher doses resulted in dose-dependent granulomatous hepatitis and histiocytic infiltration of the spleen and mesenteric lymph nodes by 1–2 weeks. B. canis was cultured from the liver, spleen, uterus, bone marrow, lung, and kidney in all groups with colonization declining at a slow but steady rate throughout the experiment. Clearance was achieved by 9 weeks 105 CFU group and by 12 weeks in the 107 CFU group, while B. canis persisted in the spleen until 12 weeks in the highest dose group. Although B. canis does not demonstrate significant replication in C57BL/6 mice, it has the ability to establish an infection, induce splenomegaly, and persist for several weeks in multiple organs. Moreover, 1 x 107 CFU appears to be a suitable challenge dose for investigating vaccine safety. [ABSTRACT FROM AUTHOR]

Based on the high sensitivity and stable fluorescence of CdTe quantum dots (QDs) in conjunction with a specific DNA aptamer, the authors describe an aptamer-based fluorescence assay for the determination of Salmonella Typhimurium. The fluorescence detection and quantification of S. Typhimurium is based on a magnetic separation system, a combination of aptamer-coated Fe3O4 magnetic particles (Apt-MNPs) and QD-labeled ssDNA2 (complementary strand of the aptamer). Apt-MNPs are employed for the specific capture of S. Typhimurium. CdTe QD-labeled ssDNA2 was used as a signaling probe. Simply, the as-prepared CdTe QD-labeled ssDNA2 was first incubated with the Apt-MNPs to form the aptamer-ssDNA2 duplex. After the addition of S. Typhimurium, they could specifically bind the DNA aptamer, leading to cleavage of the aptamer-ssDNA2 duplex, accompanied by the release of CdTe QD-labeled DNA. Thus, an increased fluorescence signal can be achieved after magnetic removal of the Apt-MNPs. The fluorescence of CdTe QDs (λexc/em = 327/612 nm) increases linearly in the concentration range of 10 to 1010 cfu•mL-1, and the limit of detection is determined to be 1 cfu•mL-1. The detection process can be performed within 2 h and is successfully applied to the analysis of spiked food samples with good recoveries from 90% to 105%. [ABSTRACT FROM AUTHOR]

Objective: To investigate the potency of a hand-held point-of-care electronic-nose to diagnose pulmonary tuberculosis (PTB) among those suspected of PTB. Methods: Setting: Lung clinics and Dr. Sardjito Hospital, Yogyakarta, Indonesia. Participants: patients with suspected PTB and healthy controls. Sampling: 5 minutes exhaled breath. Sputum-smear-microscopy, culture, chest-radiography, and follow-up for 1.5–2.5 years, were used to classify patients with suspected PTB as active PTB, probably active PTB, probably no PTB, and no PTB. After building a breath model based on active PTB, no PTB, and healthy controls (Calibration phase), we validated the model in all patients with suspected PTB (Validation phase). In each variable (sex, age, Body Mass Index, co-morbidities, smoking status, consumption of alcohol, use of antibiotics, flu symptoms, stress, food and drink intake), one stratum’s Receiver Operating Characteristic (ROC)-curve indicating sensitivity and specificity of the breath test was compared with another stratum’s ROC-curve. Differences between Area-under-the-Curve between strata (p<0.05) indicated an association between the variable and sensitivity—specificity of the breath test. Statistical analysis was performed using STATA/SE 15. Results: Of 400 enrolled participants, 73 were excluded due to extra-pulmonary TB, incomplete data, previous TB, and cancer. Calibration phase involved 182 subjects, and the result was validated in 287 subjects. Sensitivity was 85% (95%CI: 75–92%) and 78% (95%CI: 70–85%), specificity was 55% (95%CI: 44–65%) and 42% (95%CI: 34–50%), in calibration and validation phases, respectively. Test sensitivity and specificity were lower in men. Conclusion: The electronic-nose showed modest sensitivity and low specificity among patients with suspected PTB. To improve the sensitivity, a larger calibration group needs to be involved. With its portable form, it could be used for TB screening in remote rural areas and health care settings. [ABSTRACT FROM AUTHOR]

Ferlaviruses are important pathogens in snakes and other reptiles. They cause respiratory and neurological disease in infected animals and can cause severe disease outbreaks. Isolates from this genus can be divided into four genogroups–A, B, and C, as well as a more distantly related sister group, “tortoise”. Sequences from large portions (5.3 kb) of the genomes of a variety of ferlavirus isolates from genogroups A, B, and C, including the genes coding the surface glycoproteins F and HN as well as the L protein were determined and compared. In silico analyses of the glycoproteins of genogroup A, B, and C isolates were carried out. Three isolates representing these three genogroups were used in transmission studies with corn snakes (Pantherophis guttatus), and clinical signs, gross and histopathology, electronmicroscopic changes in the lungs, and isolation of bacteria from the lungs were evaluated. Analysis of the sequences supported the previous categorization of ferlaviruses into four genogroups, and criteria for definition of ferlavirus genogroups and species were established based on sequence identities (80% resp. 90%). Analysis of the ferlavirus glycoprotein models showed parallels to corresponding regions of other paramyxoviruses. The transmission studies showed clear differences in the pathogenicities of the three virus isolates used. The genogroup B isolate was the most and the group A virus the least pathogenic. Reasons for these differences were not clear based on the differences in the putative structures of their respective glycoproteins, although e.g. residue and consequential structure variation of an extended cleavage site or changes in electrostatic charges at enzyme binding sites could play a role. The presence of bacteria in the lungs of the infected animals also clearly corresponded to increased pathogenicity. This study contributes to knowledge about the structure and phylogeny of ferlaviruses and lucidly demonstrates differences in pathogenicity between strains of different genogroups. [ABSTRACT FROM AUTHOR]

Oral cholera vaccines (OCVs) are being increasingly employed, but current killed formulations generally require multiple doses and lack efficacy in young children. We recently developed a new live-attenuated OCV candidate (HaitiV) derived from a Vibrio cholerae strain isolated during the 2010 Haiti cholera epidemic. HaitiV exhibited an unexpected probiotic-like activity in infant rabbits, preventing intestinal colonization and disease by wild-type V. cholerae before the onset of adaptive immunity. However, it remained unknown whether HaitiV would behave similarly to other OCVs to stimulate adaptive immunity against V. cholerae. Here, we orally immunized adult germ-free female mice to test HaitiV’s immunogenicity. HaitiV safely and stably colonized vaccinated mice and induced known adaptive immune correlates of cholera protection within 14 days of administration. Pups born to immunized mice were protected against lethal challenges of both homologous and heterologous V. cholerae strains. Cross-fostering experiments revealed that protection was not dependent on vaccine colonization in or transmission to the pups. These findings demonstrate the protective immunogenicity of HaitiV and support its development as a new tool for limiting cholera. [ABSTRACT FROM AUTHOR]

Timely diagnosis of tuberculosis (TB) is limited in Ethiopia. We evaluated the performance of a low technology, thin layer agar, Mycobacterium tuberculosis (M.tb) culture color plate (TB-CX) test with concurrent drug susceptibility testing (DST) to isoniazid (INH), rifampin (RIF), and pyrazinamide (PZA) directly from sputum specimens. Patients undergoing examination for TB and multidrug-resistant (MDR)-TB were enrolled in Addis Ababa, Ethiopia from March 2016 to February 2017. All subjects received a GeneXpert MTB/RIF PCR test. TB-CX test results were compared to reference Löwenstein–Jensen (LJ) culture for M.tb detection and DST for susceptibility to INH and RIF. Kappa statistic was applied to test agreement between results for TB-CX test and the reference methods, a cut-off Kappa value of 0.75 was considered as high level of agreements. A total of 137 participants were analyzed: 88 (64%) were new TB cases, 49 (36%) were re-treatment cases. The TB-CX test detected M.tb and DST in an average of 13 days compared to 50 days for the conventional DST result. The sensitivity and specificity of the TB-CX test for detecting M.tb were 94% and 98%, respectively (concordance, 96%; kappa 0.91). The sensitivity of the TB-CX test to detect drug resistance to INH, RIF, and MDR-TB was 91%, 100%, and 90% respectively. The specificity of the TB-CX test for detecting INH, RIF, and MDR-TB was 94%, 40%, and 94% respectively. Overall agreement between TB-CX test and LJ DST for detection of MDR-TB was 93%. The TB-CX test showed strong agreement with the GeneXpert test for detecting M.tb (89%, kappa 0.76) but low agreement for the detection of RIF resistance (57%, kappa 0.28). The TB-CX test was found to be a good alternative method for screening of TB and selective drug resistant-TB in a timely and cost-efficient manner. [ABSTRACT FROM AUTHOR]

Background: Wolbachia’s ability to restrict arbovirus transmission makes it a promising tool to combat mosquito-transmitted diseases. Wolbachia-infected Aedes aegypti are currently being released in locations such as Brazil, which regularly experience concurrent outbreaks of different arboviruses. A. aegypti can become co-infected with, and transmit multiple arboviruses with one bite, which can complicate patient diagnosis and treatment. Methodology/principle findings: Using experimental oral infection of A. aegypti and then RT-qPCR, we examined ZIKV/DENV-1 and ZIKV/DENV-3 co-infection in Wolbachia-infected A. aegypti and observed that Wolbachia-infected mosquitoes experienced lower prevalence of infection and viral load than wildtype mosquitoes, even with an extra infecting virus. Critically, ZIKV/DENV co-infection had no significant impact on Wolbachia’s ability to reduce viral transmission. Wolbachia infection also strongly altered expression levels of key immune genes Defensin C and Transferrin 1, in a virus-dependent manner. Conclusions/significance: Our results suggest that pathogen interference in Wolbachia-infected A. aegypti is not adversely affected by ZIKV/DENV co-infection, which suggests that Wolbachia-infected A. aegypti will likely prove suitable for controlling mosquito-borne diseases in environments with complex patterns of arbovirus transmission. [ABSTRACT FROM AUTHOR]

Immunoglobulin A is a class of antibodies produced by the adaptive immune system and secreted into the gut lumen to fight pathogenic bacteria. We recently demonstrated that the main physical effect of these antibodies is to enchain daughter bacteria, i.e. to cross-link bacteria into clusters as they divide, preventing them from interacting with epithelial cells, thus protecting the host. These links between bacteria may break over time. We study several models using analytical and numerical calculations. We obtain the resulting distribution of chain sizes, that we compare with experimental data. We study the rate of increase in the number of free bacteria as a function of the replication rate of bacteria. Our models show robustly that at higher replication rates, bacteria replicate before the link between daughter bacteria breaks, leading to growing cluster sizes. On the contrary at low growth rates two daughter bacteria have a high probability to break apart. Thus the gut could produce IgA against all the bacteria it has encountered, but the most affected bacteria would be the fast replicating ones, that are more likely to destabilize the microbiota. Linking the effect of the immune effectors (here the clustering) with a property directly relevant to the potential bacterial pathogeneicity (here the replication rate) could avoid to make complex decisions about which bacteria to produce effectors against. [ABSTRACT FROM AUTHOR]

Background: Assessing Mycobacterium tuberculosis (TB) viability by fluorescein diacetate (FDA) microscopy can predict TB culture results, treatment response and infectiousness. However, diverse methods have been published. We aimed to optimise FDA microscopy, minimising sputum processing, biohazard and complexity for use in resource-constrained settings. Methods and results: Optimization: Patients with smear-positive pulmonary TB before treatment and healthy control participants provided sputa. These were divided into equal aliquots that were tested directly or after NaOH centrifuge-decontamination. Each aliquot was cultured and used to prepare slides (n = 80). FDA microscopy used: 1 or 3 drops of sputum; with/out acid-alcohol wash; with/out phenol sterilization; with 0/30/60 seconds KMnO4 quenching. Control samples all had negative culture and microscopy results. FDA microscopy had higher sensitivity when performed directly (without centrifuge-decontamination) on 1 drop of sputum (P<0.001), because 3 drops obscured microscopy. Acid-alcohol wash and KMnO4 quenching made bacilli easier to identity (P = 0.005). Phenol sterilization did not impair microscopy (P>0.1). Validation: The 2 protocols that performed best in the optimization experiments were reassessed operationally by comparing duplicate slides (n = 412) stained with KMnO4 quenching for 30 versus 60 seconds. FDA microscopy results were similar (P = 0.4) and highly reproducible, with 97% of counts agreeing within +/-1 logarithm. Storage: Smear microscopy slides and aliquots of the sputum from which they were made were stored for 4 weeks. Twice-weekly, paired slides (n = 80) were stained with freshly prepared versus stored FDA and read quantitatively. Storing sputum, microscopy slides or FDA solution at 4°C or room temperature had no effect on FDA microscopy results (all P>0.2). Cost: Material costs for each slide tested by FDA microscopy using reagents purchased locally were USD $0.05 and required the same equipment, time and skills as auramine acid-fast microscopy. Conclusions: We recommend a simple, bio-secure protocol for FDA microscopy that provides sensitive and repeatable results without requiring centrifugation. [ABSTRACT FROM AUTHOR]

Background: A nonsputum blood test capable of predicting progression of healthy individuals to active tuberculosis (TB) before clinical symptoms manifest would allow targeted treatment to curb transmission. We aimed to develop a proteomic biomarker of risk of TB progression for ultimate translation into a point-of-care diagnostic.Methods and Findings: Proteomic TB risk signatures were discovered in a longitudinal cohort of 6,363 Mycobacterium tuberculosis-infected, HIV-negative South African adolescents aged 12-18 years (68% female) who participated in the Adolescent Cohort Study (ACS) between July 6, 2005 and April 23, 2007, through either active (every 6 months) or passive follow-up over 2 years. Forty-six individuals developed microbiologically confirmed TB disease within 2 years of follow-up and were selected as progressors; 106 nonprogressors, who remained healthy, were matched to progressors. Over 3,000 human proteins were quantified in plasma with a highly multiplexed proteomic assay (SOMAscan). Three hundred sixty-one proteins of differential abundance between progressors and nonprogressors were identified. A 5-protein signature, TB Risk Model 5 (TRM5), was discovered in the ACS training set and verified by blind prediction in the ACS test set. Poor performance on samples 13-24 months before TB diagnosis motivated discovery of a second 3-protein signature, 3-protein pair-ratio (3PR) developed using an orthogonal strategy on the full ACS subcohort. Prognostic performance of both signatures was validated in an independent cohort of 1,948 HIV-negative household TB contacts from The Gambia (aged 15-60 years, 66% female), longitudinally followed up for 2 years between March 5, 2007 and October 21, 2010, sampled at baseline, month 6, and month 18. Amongst these contacts, 34 individuals progressed to microbiologically confirmed TB disease and were included as progressors, and 115 nonprogressors were included as controls. Prognostic performance of the TRM5 signature in the ACS training set was excellent within 6 months of TB diagnosis (area under the receiver operating characteristic curve [AUC] 0.96 [95% confidence interval, 0.93-0.99]) and 6-12 months (AUC 0.76 [0.65-0.87]) before TB diagnosis. TRM5 validated with an AUC of 0.66 (0.56-0.75) within 1 year of TB diagnosis in the Gambian validation cohort. The 3PR signature yielded an AUC of 0.89 (0.84-0.95) within 6 months of TB diagnosis and 0.72 (0.64-0.81) 7-12 months before TB diagnosis in the entire South African discovery cohort and validated with an AUC of 0.65 (0.55-0.75) within 1 year of TB diagnosis in the Gambian validation cohort. Signature validation may have been limited by a systematic shift in signal magnitudes generated by differences between the validation assay when compared to the discovery assay. Further validation, especially in cohorts from non-African countries, is necessary to determine how generalizable signature performance is.Conclusions: Both proteomic TB risk signatures predicted progression to incident TB within a year of diagnosis. To our knowledge, these are the first validated prognostic proteomic signatures. Neither meet the minimum criteria as defined in the WHO Target Product Profile for a progression test. More work is required to develop such a test for practical identification of individuals for investigation of incipient, subclinical, or active TB disease for appropriate treatment and care. [ABSTRACT FROM AUTHOR]

Leptospirosis is a zoonosis with a worldwide distribution, caused by pathogenic spirochetes of the genus Leptospira. The classification and identification of leptospires can be conducted by both genotyping and serotyping which are time-consuming and established in few reference laboratories. This study used matrix assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) as rapid and accurate tool for the identification of leptospires. The whole cell protein spectra of 116 Leptospira isolates including 15 references Leptospira spp. (pathogenic, n = 8; intermediate, n = 2; non-pathogenic, n = 5) and 101 Leptospira spp. clinical isolates was created as an in-house MALDI-TOF MS database. Ninety-seven clinical isolates from Thailand and Laos was validated with these protein spectra and revealed 98.9% correct identification when compared with 16S rRNA gene sequences method. Moreover, MALDI-TOF MS could identify spiked leptospires whole cell in urine. Biomarkers for differentiation of leptospires phylogeny and specific protein spectra for most found Leptospira spp. in this area (L. interrogans, L. kirschneri, L. borgpetersenii) based on MALDI-MS algorithm were demonstrated. [ABSTRACT FROM AUTHOR]

No aetiology is found in up to 40% of men with symptomatic urethritis. Male partners of women with bacterial vaginosis (BV) may be at higher risk of non-gonococcal urethritis (NGU). The aim of this study was to examine the role of BV associated bacteria in first-void urine (FVU) in 97 asymptomatic men without urethritis (controls) and 44 men (cases) with NGU including 20 men with idiopathic urethritis (IU) attending a Swedish STD-clinic between January and October 2010. BV-associated bacteria and ureaplasmas were detected by quantitative PCR assays. All BV associated bacteria, except Megasphaera-like type 1, were strongly positively correlated with U. urealyticum p<0.005 and even stronger with the combined U. urealyticum and U. parvum load (p<0.0005) suggesting that ureaplasma induced elevated pH may stimulate the growth of BV associated bacteria. No statistically significant differences were found between IU cases and controls in the prevalence or load of BV associated bacteria or ureaplasmas. In multiple logistic regression, Megasphaera-like type 1 was associated with IU (p = 0.03), but most positive FVU samples contained very few bacteria and the finding may not be clinically relevant. [ABSTRACT FROM AUTHOR]

Introduction: Gonorrhea (GC) infection caused by Neisseria gonorrhoeae has been steadily increasing in Thailand over the last decade. Men who have sex with men (MSM) are at high risk for gonorrhea infection. Materials and methods: In this study, we determined the prevalence of and risk factors associated with gonococcal infections by three anatomical sites among MSM. We have conducted a cross-sectional analysis of a sexually transmitted disease (STD), gonorrhea among MSM attending two STD clinics in Khon Kaen, Thailand. We included 358 MSM over 18 years of age. Data were collected using self-administered questionnaire. In each participant, an oropharyngeal, anorectal, and endourethral swab were tested with culture and nucleic acid amplification test (NAAT). However, 267 urine samples were tested by both methods. Factors associated with gonorrhea infections were assessed using univariate and multivariate logistic regression. Results: One hundred and ninety-five out of 358 (54.47%) MSM tested were found to be positive for gonorrhea using a porA gene targeted NAAT by Real-time PCR with TaqMan probes, but there was no positive result by culture. The gonorrheal prevalence for male genital site, anal, and oropharyngeal, were 34.73% (95%CI 33.07, 45.08), 29.01% (95%CI 24.61, 34.33), and 27.93% (95%CI 23.35, 32.89), respectively, while 5.9% (21/355) were positive for gonococcal infection in all anatomic sites (oropharynx + anus + urethra) of one participant. Previous history of diagnosed STDs was a significant factor associated urethral gonorrhea (odds ratio = 3.52, 95%CI 1.87–6.66, P Value< 0.001). In addition, having more than one partner was increased urethral gonorrhea (adjusted odds ratio = 2.26, 95%CI 1.10–4.68, P Value = 0.026). 100% of condom use was found decreasing urethral infection (adjusted odds ratio = 0.39, 95%CI 0.15–0.99, P Value = 0.046). Conclusions: The most common anatomic site of gonorrhea infection was male genital site, and the independent risk factors were having history of diagnosed STDs and having more than one partner in the past 3 months, but 100% condom use was a protective factor of this infection. [ABSTRACT FROM AUTHOR]

The article discusses about approaches towards the diagnosis and treatment of leprosy in Cambodia. It looks at the retrospective active case finding (RACF) program in the country for tracing leprosy patients, screening of the patients' contacts for leprosy, and providing a single dose of rifampicin (SDR) as post exposure prophylaxis (PEP) to the contacts.

Background: Timely diagnosis of tuberculosis disease is critical for positive patient outcomes, yet potentially millions go undiagnosed or unreported each year. Sputum is widely used as the testing input, but limited by its complexity, heterogeneity, and sourcing problems. Finding methods to interrogate noninvasive, non-sputum clinical specimens is indispensable to improving access to tuberculosis diagnosis and care. In this work, economical plasmonic gratings were used to analyze tuberculosis biomarker lipoarabinomannan (LAM) from clinical urine samples by single molecule fluorescence assay (FLISA) and compared with gold standard sputum GeneXpert MTB/ RIF, culture, and reference ELISA testing results. Methods and Findings: In this study, twenty sputum and urine sample sets were selected retrospectively from a repository of HIV-negative patient samples collected before initiation of anti-tuberculosis therapy. GeneXpert MTB/RIF and culture testing of patient sputum confirmed the presence or absence of pulmonary tuberculosis while all patient urines were reference ELISA LAM-negative. Plasmonic gratings produced by low-cost soft lithography were bound with anti-LAM capture antibody, incubated with patient urine samples, and biotinylated detection antibody. Fluorescently labeled streptavidin revealed single molecule emission by epifluorescence microscope. Using a 1 fg/mL baseline for limit of detection, single molecule FLISA demonstrated good qualitative agreement with gold standard tests on 19 of 20 patients, including accurately predicting the gold-standard-negative patients, while one gold-standard-positive patient produced no observable LAM in urine. Conclusions: Single molecule FLISA by plasmonic grating demonstrated the ability to quantify tuberculosis LAM from complex urine samples of patients from a high endemic setting with negligible interference from the complex media itself. Moreover, agreement with patient diagnoses by gold standard testing suggests that single molecule FLISA could be used as a highly sensitive test to diagnose tuberculosis noninvasively. [ABSTRACT FROM AUTHOR]

Purpose: Homeless persons have a high risk for tuberculosis. The prevalence of latent tuberculosis infection and the risk for a progression to active tuberculosis is higher in the homeless than in the general population. The objective was to assess the prevalence and risk factors of tuberculosis/latent tuberculosis infection in a homeless population in Germany. Methods: Homeless individuals (n = 150) were enrolled in a cross-sectional study at three shelters in Münster, Germany (October 2017–July 2018). All participants were screened using an ELISPOT interferon-γ release assay (IGRA). Those participants tested positive/borderline by IGRA provided three sputa for microbiological analysis (line probe assay, microscopy, culture) and underwent a chest X-ray to screen for active pulmonary TB. Risk factors for tuberculosis/latent tuberculosis infection were analysed using a standardized questionnaire. Results: Of the 142 evaluable IGRA, 21 (15%) were positive and two (1%) were borderline. No participant with a positive/borderline IGRA had an active tuberculosis as assessed by chest X-ray and microbiology. A negative IGRA was associated with a citizenship of a low-incidence country for tuberculosis (according to WHO, p = 0.01), low-incidence country of birth (p<0.001) or main residence in a low-incidence country in the past five years (p = 0.002). Conclusions: The prevalence of latent tuberculosis infection (diagnosed by a positive/borderline IGRA) was 16%; no active tuberculosis was detected. The highest risk for latent tuberculosis infection was found in patients from high-incidence countries. This population at risk should be either treated for latent tuberculosis infection or need to be monitored to early detect a progression into active disease. [ABSTRACT FROM AUTHOR]

Background: Helminth infections are known to regulate cytokine responses in both pulmonary and latent tuberculosis infection. Whether helminth infections also modulate cytokine responses in extra-pulmonary tuberculosis, specifically tuberculous lymphadenitis (TBL), has not been examined thus far. Methodology: Hence, to determine the cytokine profile in helminth-TBL coinfection, we measured the systemic and mycobacterial (TB)–antigen stimulated levels of Type 1, Type 2, Type 17, regulatory and pro-inflammatory cytokines in TBL individuals coinfected with or without Strongyloides stercoralis (Ss) infection. Significant findings: TBL-Ss+ individuals have significantly higher bacterial burdens in the affected lymph nodes in comparison to TBL-Ss- individuals. TBL-Ss+ individuals exhibit significantly enhanced plasma levels of Type 2 (IL-5 and IL-13), Type 17 (IL-17 and IL-22) and regulatory (IL-10) cytokines in comparison to TBL-Ss- individuals. In contrast, TBL-Ss+ individuals exhibit significantly diminished plasma levels of pro-inflammatory cytokines (IL-1α and GM-CSF) in comparison to TBL-Ss- individuals. TBL-Ss+ individuals also exhibit significantly diminished unstimulated or mycobacterial—antigen stimulated levels of Type 1, Type 17 or IL-1 family cytokines in comparison to TBL-Ss- individuals but no differences in mitogen stimulated cytokine levels. Conclusion: Therefore, our data reveal a profound influence of Ss infection on the bacteriological profile of TBL and suggesting that the underlying modulation of cytokine responses might be a mechanism by which this helminth infection could impart a detrimental effect on the pathogenesis of TBL disease. [ABSTRACT FROM AUTHOR]

Leprosy is a chronic infection where the skin and peripheral nervous system is invaded by Mycobacterium leprae. The infection mechanism remains unknown in part because culture methods have not been established yet for M. leprae. Mce1A protein (442 aa) is coded by mce1A (1326 bp) of M. leprae. The Mce1A homolog in Mycobacterium tuberculosis is known to be associated with M. tuberculosis epithelial cell entry, and survival and multiplication within macrophages. Studies using recombinant proteins have indicated that Mce1A of M. leprae is also associated with epithelial cell entry. This study is aimed at identifying particular sequences within Mce1A associated with M. leprae epithelial cell entry. Recombinant proteins having N-terminus and C-terminus truncations of the Mce1A region of M. leprae were created in Escherichia coli. Entry activity of latex beads, coated with these truncated proteins (r-lep37 kDa and r-lep27 kDa), into HeLa cells was observed by electron microscopy. The entry activity was preserved even when 315 bp (105 aa) and 922 bp (308 aa) was truncated from the N-terminus and C-terminus, respectively. This 316–921 bp region was divided into three sub-regions: 316–531 bp (InvX), 532–753 bp (InvY), and 754–921 bp (InvZ). Each sub-region was cloned into an AIDA vector and expressed on the surface of E. coli. Entry of these E. coli into monolayer-cultured HeLa and RPMI2650 cells was observed by electron microscopy. Only E. coli harboring the InvX sub-region exhibited cell entry. InvX was further divided into 4 domains, InvXa—InvXd, containing sequences 1–24 aa, 25–46 aa, 47–57 aa, and 58–72 aa, respectively. Recombinant E. coli, expressing each of InvXa—InvXd on the surface, were treated with antibodies against these domains, then added to monolayer cultured RPMI cells. The effectiveness of these antibodies in preventing cell entry was studied by colony counting. Entry activity was suppressed by antibodies against InvXa, InvXb, and InvXd. This suggests that these three InvX domains of Mce1A are important for M. leprae invasion into nasal epithelial cells. [ABSTRACT FROM AUTHOR]

Serum vibriocidal antibody assays have long been used to evaluate the immunogenicity of cholera vaccines formulated with killed whole-cell Vibrio cholerae. However, the antibody isotypes responsible for the serum vibriocidal activity are not fully characterized. In this study, we examined 20 clinical serum samples obtained from human subjects who had been vaccinated with a killed, whole-cell cholera vaccine and a positive control, human convalescent sera with high vibriocidal activity, to determine which isotype antibody is associated with the vibriocidal activity. Antibody isotypes from pooled convalescent sera were fractionated by size-exclusion column chromatography, and the major vibriocidal activity was detected in the IgM fraction. Depletion of IgM antibodies in the convalescent sera produced a significant (P<0.05) decrease in vibriocidal activity (16-fold decrease), whereas only a small change was observed with depletion of IgG or IgA. In addition, anti-LPS IgM antibody showed the highest correlation with vibriocidal activity (Spearman correlation coefficient r = 0.846) among antibody isotypes against heat-killed V. cholerae, lipopolysaccharide (LPS), or major outer membrane protein (Omp U), while total IgG, IgA, or IgM antibody level was not correlated with vibriocidal activity in the 20 human clinical serum samples. Furthermore, human convalescent sera significantly (P<0.001) inhibited the attachment of V. cholerae to HT-29, a human intestinal epithelial cell in vitro. Interestingly, IgM-depleted convalescent sera could not effectively inhibit bacterial adherence compared with non-depleted sera (P<0.05). Finally, bacterial adhesion was significantly inhibited by sera with high vibriocidal titer compared with low-titer sera (P = 0.014). Collectively, we demonstrated that anti-V. cholerae LPS IgM is highly correlated with serum vibriocidal activity and it could be a surrogate antibody isotype representing protective antibodies against V. cholerae. [ABSTRACT FROM AUTHOR]

FBXO7 encodes an F box containing protein that interacts with multiple partners to facilitate numerous cellular processes and has a canonical role as part of an SCF E3 ubiquitin ligase complex. Mutation of FBXO7 is responsible for an early onset Parkinsonian pyramidal syndrome and genome-wide association studies have linked variants in FBXO7 to erythroid traits. A putative orthologue in Drosophila, nutcracker, has been shown to regulate the proteasome, and deficiency of nutcracker results in male infertility. Therefore, we reasoned that modulating Fbxo7 levels in a murine model could provide insights into the role of this protein in mammals. We used a targeted gene trap model which retained 4–16% residual gene expression and assessed the sensitivity of phenotypic traits to gene dosage. Fbxo7 hypomorphs showed regenerative anaemia associated with a shorter erythrocyte half-life, and male mice were infertile. Alterations to T cell phenotypes were also observed, which intriguingly were both T cell intrinsic and extrinsic. Hypomorphic mice were also sensitive to infection with Salmonella, succumbing to a normally sublethal challenge. Despite these phenotypes, Fbxo7 hypomorphs were produced at a normal Mendelian ratio with a normal lifespan and no evidence of neurological symptoms. These data suggest that erythrocyte survival, T cell development and spermatogenesis are particularly sensitive to Fbxo7 gene dosage. [ABSTRACT FROM AUTHOR]

Background: Although QuantiFERON-TB Gold Plus (QFT-Plus), a new interferon-gamma release assay, has shown good performance in adults, little data is available in children. Methods: De-identified data from TB-suspected patients age <18 years with QFT-Plus results, who were admitted or screened at the National Lung Hospital (NLH) in Ha Noi, Vietnam in 2017, were assessed. Logistic regression analyses were performed to determine the characteristics associated with having a positive QFT-Plus result. Sensitivity, both overall and in subgroups of pulmonary TB only (PTB), extra-pulmonary TB (EPTB) only, and both PTB and EPTB were calculated. Results: Of 222 children with available QFT-Plus results, 33 were classified as confirmed TB, of whom 18 had QFT-Plus (+) and 15 had QFT-Plus (-). Multiple logistic regression modeling suggested that age, history of TB, and confirmed TB were significantly associated with having a positive QFT-Plus result with an area under the ROC curve of 0.77. QFT-Plus sensitivity in PTB only, EPTB, and both PTB and EPTB patients was 84.2%, 14.3% and 14.3%, respectively. The overall sensitivity of the QFT-Plus assay (regardless PTB or EPTB) in children was 54.5%. Conclusion: Although QFT-Plus had a good sensitivity in children having exclusive PTB, it had poor sensitivity in EPTB. [ABSTRACT FROM AUTHOR]

Tsetse flies (Glossina spp.) vector pathogenic trypanosomes (Trypanosoma spp.) in sub-Saharan Africa. These parasites cause human and animal African trypanosomiases, which are debilitating diseases that inflict an enormous socio-economic burden on inhabitants of endemic regions. Current disease control strategies rely primarily on treating infected animals and reducing tsetse population densities. However, relevant programs are costly, labor intensive and difficult to sustain. As such, novel strategies aimed at reducing tsetse vector competence require development. Herein we investigated whether Kosakonia cowanii Zambiae (Kco_Z), which confers Anopheles gambiae with resistance to Plasmodium, is able to colonize tsetse and induce a trypanosome refractory phenotype in the fly. Kco_Z established stable infections in tsetse’s gut and exhibited no adverse effect on the fly’s survival. Flies with established Kco_Z infections in their gut were significantly more refractory to infection with two distinct trypanosome species (T. congolense, 6% infection; T. brucei, 32% infection) than were age-matched flies that did not house the exogenous bacterium (T. congolense, 36% infected; T. brucei, 70% infected). Additionally, 52% of Kco_Z colonized tsetse survived infection with entomopathogenic Serratia marcescens, compared with only 9% of their wild-type counterparts. These parasite and pathogen refractory phenotypes result from the fact that Kco_Z acidifies tsetse’s midgut environment, which inhibits trypanosome and Serratia growth and thus infection establishment. Finally, we determined that Kco_Z infection does not impact the fecundity of male or female tsetse, nor the ability of male flies to compete with their wild-type counterparts for mates. We propose that Kco_Z could be used as one component of an integrated strategy aimed at reducing the ability of tsetse to transmit pathogenic trypanosomes. [ABSTRACT FROM AUTHOR]

Setting: Community based tuberculosis (TB) prevalence surveys in ten sites across India during 2006–2012 Objective: To re-analyze data of recent sub-national surveys using uniform statistical methods and obtain a pooled national level estimate of prevalence of TB. Methods: Individuals ≥15 years old were screened by interview for symptoms suggestive of Pulmonary TB (PTB) and history of anti-TB treatment; additional screening by chest radiography was undertaken in five sites. Two sputum specimens were examined by smear and culture among Screen-positives. Prevalence in each site was estimated after imputing missing values to correct for bias introduced by incompleteness of data. In five sites, prevalence was corrected for non-screening by radiography. Pooled prevalence of bacteriologically positive PTB was estimated using Random Effects Model after excluding data from one site. Overall prevalence of TB (all ages, all types) was estimated by adjusting for extra-pulmonary TB and Pediatric TB. Results: Of 769290 individuals registered, 715989 were screened by interview and 294532 also by radiography. Sputum specimen were examined from 50 852 individuals. Estimated prevalence of smear positive, culture positive and bacteriologically positive PTB varied between 108.4–428.1, 147.9–429.8 and 170.8–528.4 per 100000 populations in different sites. Pooled estimate of prevalence of bacteriologically positive PTB was 350.0 (260.7, 439.0). Overall prevalence of TB was estimated at 300.7 (223.7–377.5) in 2009, the mid-year of surveys. Prevalence was significantly higher in rural compared to urban areas. Conclusion: TB burden continues to be high in India suggesting further strengthening of TB control activities. [ABSTRACT FROM AUTHOR]

Molecular typing techniques are useful in understanding tuberculosis epidemiology; yet, they have been under-utilised at the human-animal interface in Nigeria. Sixty-four Mycobacterium tuberculosis complex (MTBC) isolates including 42 M. tuberculosis, 13 M. bovis and nine M. africanum obtained from livestock workers (LW, n = 47) and their cattle (n = 17) in three geographical zones of Nigeria were genotyped to identify and evaluate the genetic diversity of the circulating MTBC using spoligotyping. Distribution into clades of M. tuberculosis revealed; 45.3% Uganda I- [SIT46- cattle: 1; LW: 28], 14.1% Latin American Mediterranean- [SIT61, cattle: 1; LW: 8], and 1.6% T- [SIT53—LW: 1]. The M. bovis strains were 6.3% SB0944 [cattle: 4] and 1.6% each of SB0300, SB1026, SB1027 and SB1439 [cattle: 4]. Seventeen MTBC isolates [cattle: 7; LW: 10] yielded 14 new spoligotype patterns including three M. tuberculosis strains (three isolates), five M. bovis strains (five isolates) and six M. africanum strains (nine isolates), two of which belonged to MAF1. Only few families namely, the not previously described Uganda I-, LAM and SB0944 are predominant among the LW and cattle, with other types in lower prevalences. The strain population structure indicates an intriguing diversity and possible zoonotic linkage with consequences for TB control in the country. The need to employ newer molecular techniques such as Mycobacterial Interspersed Repetitive Unit-Variable Number Tandem Repeats and whole genome sequence to decipher circulating MTBC strains in Nigeria is advocated. [ABSTRACT FROM AUTHOR]

Objectives: To examine whether post-treatment Lyme disease syndrome (PTLDS) defined by fatigue, cognitive complaints and widespread pain following the treatment of Lyme disease is associated with small fiber neuropathy (SFN) manifesting as autonomic and sensory dysfunction. Methods: This single center, retrospective study evaluated subjects with PTLDS. Skin biopsies for assessment of epidermal nerve fiber density (ENFD), sweat gland nerve fiber density (SGNFD) and functional autonomic testing (deep breathing, Valsalva maneuver and tilt test) were performed to assess SFN, severity of dysautonomia and cerebral blood flow abnormalities. Heart rate, end tidal CO2, blood pressure, and cerebral blood flow velocity (CBFv) from middle cerebral artery using transcranial Doppler were monitored. Results: 10 participants, 5/5 women/men, age 51.3 ± 14.7 years, BMI 27.6 ± 7.3 were analyzed. All participants were positive for Lyme infection by CDC criteria. At least one skin biopsy was abnormal in all ten participants. Abnormal ENFD was found in 9 participants, abnormal SGNFD in 5 participants, and both abnormal ENFD and SGNFD were detected in 4 participants. Parasympathetic failure was found in 7 participants and mild or moderate sympathetic adrenergic failure in all participants. Abnormal total CBFv score was found in all ten participants. Low orthostatic CBFv was found in 7 participants, three additional participants had abnormally reduced supine CBFv. Conclusions: SFN appears to be associated with PTLDS and may be responsible for certain sensory symptoms. In addition, dysautonomia related to SFN and abnormal CBFv also seem to be linked to PTLDS. Reduced orthostatic CBFv can be associated with cerebral hypoperfusion and may lead to cognitive dysfunction. Autonomic failure detected in PTLDS is mild to moderate. SFN evaluation may be useful in PTLDS. [ABSTRACT FROM AUTHOR]