Risk factors for persistent abnormality on chest radiographs at 12-weeks post hospitalisation with PCR confirmed COVID-19.

BACKGROUND: The long-term consequences of COVID-19 remain unclear. There is concern a proportion of patients will progress to develop pulmonary fibrosis. We aimed to assess the temporal change in CXR infiltrates in a cohort of patients following hospitalisation for COVID-19. METHODS: We conducted a single-centre prospective cohort study of patients admitted to University Hospital Southampton with confirmed SARS-CoV2 infection between 20th March and 3rd June 2020. Patients were approached for standard-of-care follow-up 12-weeks after hospitalisation. Inpatient and follow-up CXRs were scored by the assessing clinician for extent of pulmonary infiltrates; 0-4 per lung (Nil = 0, < 25% = 1, 25-50% = 2, 51-75% = 3, > 75% = 4). RESULTS: 101 patients with paired CXRs were included. Demographics: 53% male with a median (IQR) age 53.0 (45-63) years and length of stay 9 (5-17.5) days. The median CXR follow-up interval was 82 (77-86) days with median baseline and follow-up CXR scores of 4.0 (3-5) and 0.0 (0-1) respectively. 32% of patients had persistent CXR abnormality at 12-weeks. In multivariate analysis length of stay (LOS), smoking-status and obesity were identified as independent risk factors for persistent CXR abnormality. Serum LDH was significantly higher at baseline and at follow-up in patients with CXR abnormalities compared to those with resolution. A 5-point composite risk score (1-point each; LOS ≥ 15 days, Level 2/3 admission, LDH > 750 U/L, obesity and smoking-status) strongly predicted risk of persistent radiograph abnormality (0.81). CONCLUSION: Persistent CXR abnormality 12-weeks post COVID-19 was common in this cohort. LOS, obesity, increased serum LDH, and smoking-status were risk factors for radiograph abnormality. These findings require further prospective validation.

Evidence of mercury biomagnification in the food chain of the cardinal tetra Paracheirodon axelrodi (Osteichthyes: Characidae) in the Rio Negro, central Amazon, Brazil.

In this study, nitrogen stable isotope (δ(15) N) and total mercury (THg) analyses were conducted on algae, submersed and emergent macrophytes, shrubs and trees, Macrobrachium sp. and Paracheirodon axelrodi collected in three streams that drain a large interfluvial region in the middle Rio Negro, Amazonas State, Brazil. Samples were collected during different hydrological periods over 12 months in lower stream reaches and their headwaters; the latter being characterized by shallow, open-canopy swamps. Additionally, δ(15) N values and mercury concentrations of Paracheirodon simulans and Cichla spp. from the middle Rio Negro were analysed to demonstrate THg biomagnification in the food web. The highest mercury levels of P. axelrodi were found in small individuals, which were collected principally in the low water period. The log10 THg-δ(15) N relationship of vascular plants and algae, Macrobrachium sp., Paracheirodon spp. and Cichla spp. showed significant mercury biomagnification among trophic levels, with regression slopes of 0·15 and 0·25 for the entire food web and heterotrophs-only food web, respectively. The mean ± s.d. THg concentrations for Macrobrachium sp., P. axelrodi, P. simulans and Cichla spp. were 63·6 ± 23·7, 104·5 ± 40·0, 112·3 ± 31·4 and 418·5 ± 188·1 ng g(-1) wet mass, respectively. Elevated levels of mercury found in Paracheirodon spp. and top predators such as Cichla spp. in a remote area far from anthropogenic inputs provide evidence that high mercury concentrations occur naturally in Rio Negro aquatic food webs.

Pulmonary infection with Scedosporium prolificans in an immunocompetent individual.

We report a case of histologically confirmed Scedosporium prolificans pulmonary infection secondary to long-standing bronchiectasis that necessitated excision lung surgery. This case emphasizes the difficulties with the medical management of deep-seated fungal infections.

Enhanced antimycobacterial response to recombinant Mycobacterium bovis BCG expressing latency-associated peptide.

With a view to exploring the role of transforming growth factor beta (TGF-beta) during mycobacterial infection, recombinant clones of bacillus Calmette-Guérin (BCG) were engineered to express the natural antagonist of TGF-beta, latency-activated peptide (LAP). Induction of TGF-beta activity was reduced when macrophages were infected with BCG expressing the LAP construct (LAP-BCG). There was a significant reduction in the growth of LAP-BCG in comparison to that of control BCG following intravenous infection in a mouse model. The enhanced control of mycobacterial replication was associated with an increase in the production of gamma interferon by splenocytes challenged during the acute stage of infection but with a diminished recall response assessed after 13 weeks. Organ weight and hydroxyproline content, representing tissue pathology, were also lower in mice infected with LAP-BCG. The results are consistent with the hypothesis that TGF-beta has a detrimental effect on mycobacterial immunity. While a reduction in TGF-beta activity augments the initial response to BCG vaccination, early bacterial clearance may adversely affect the induction of a long-term memory response by LAP-BCG.

Bronchopulmonary and mediastinal leishmaniasis: an unusual clinical presentation of Leishmania donovani infection.

We describe a case of unusual leishmaniasis in a Sudanese man with a history of progressively enlarging granulomatous mediastinal lymphadenopathy, worsening hemoptysis, and an intense mucosal granulomatous inflammatory response in the large bronchi. Leishmania donovani DNA was detected in bronchial biopsies by polymerase chain reaction. This is a novel description of human leishmanial infection in an immunocompetent patient involving this anatomical site. The patient's condition improved clinically, spirometrically, and radiologically after a course of treatment with amphotericin B. The cell-mediated immune response was analyzed before, during, and after successful antileishmanial chemotherapy.

Tumour necrosis factor-alpha production in human alveolar macrophages: modulation by inhaled corticosteroid.

Using an ex vivo alveolar macrophage model, the hypothesis that inhaled preparations of corticosteroids might have important anti-inflammatory effects on cells of the peripheral airway was tested. The tumour necrosis factor (TNF)-alpha-inducing potential of three glycolipid preparations from nonpathogenic (arabinofuranasyl lipoarabinomannan (LAM (Ara-LAM)) and virulent (mannase LAM (ManLAM)) mycobacteria and Gram-negative bacteria (lipopolysaccharide (LPS)), in primary alveolar macrophage preparations was investigated. A novel inhaled chlorofluorocarbon (CFC)-free preparation of beclomethasone dipropionate (hydrofluoroalkane 134a (HFA)-BDP) with increased peripheral lung deposition was investigated for its ability to modulate glycolipidinduced TNF-alpha production by human alveolar macrophages, in comparison with a CFC-containing preparation and placebo. Compared to the basal TNF-alpha bioactivity of 0.72 ng x mL-1 (geometric mean), the TNF-alpha bioactivity in the macrophage preparation increased following incubation with LPS (138 ng x mL-1, p<0.001), AraLAM (12.6 ng-mL-1, p<0.001) and ManLAM (1.42 ng x mL-1, p=0.02). HFA-BDP, administered in vivo, significantly reduced LPS- and ManLAM-induced TNF-alpha production by alveolar macrophages cultured ex vivo. No change in glycolipid-induced TNF-alpha production was observed following in vivo administration of CFC-BDP or HFA-placebo. This is the first demonstration of an immunomodulatory effect on alveolar cells of corticosteroid delivered via metered dose inhaler. The present findings suggest that alveolar deposition of beclomethasone dipropionate is capable of modulating the inflammatory potential of the alveolar macrophage population.

Bacille Calmette-Guérin (BCG)-associated inflammation and fibrosis: modulation by recombinant BCG expressing interferon-gamma (IFN-gamma).

Immunization with existing BCG vaccines has failed to confer consistent protection against tuberculosis. One of the ways to improve the efficacy of BCG is by enhancing its ability to induce a type-1 T cell response. However, this approach carries the risk that enhanced immunoreactivity may exacerbate tissue pathology associated with vaccination. The aim of the present study was to determine whether use of a recombinant BCG expressing IFN-gamma (BCG-IFN) would result in an alteration in the pattern of inflammation and local tissue fibrosis. A murine intravenous BCG infection model was used in which there was a time- and dose-dependent increase in the weight and number of granulomas in the liver. Infection was associated with increased inflammatory activity in the liver, as shown by the increase in expression of inducible nitric oxide synthase (iNOS) assessed by immunochemistry and by measurement of specific mRNA, and in fibrosis measured by hydroxyproline content of the liver and percentage of granuloma cells staining positively for type 1 procollagen. Infection with BCG-IFN resulted in a reduction in organ weight and bacterial load on day 21 compared with infection with control BCG transformed with vector alone (BCG-plasmid). By day 21, there was also a reduction in iNOS mRNA and iNOS+ cells in granulomas in mice infected with BCG-IFN compared with infection with BCG-plasmid, and a similar reduction in both total number of granulomas and liver hydroxyproline content. These results demonstrate that the granulomas in the areas of mycobacterial infection are active sites of both inflammation and fibrosis, and that the local expression of IFN-gamma by the recombinant BCG results in more efficient bacterial clearance which is accompanied by a reduction in tissue pathology.

HIV and tuberculosis co-infection in an inner London hospital--a prospective anonymized seroprevalence study.

OBJECTIVES: Since 1987 there has been an increase in tuberculosis notifications in the U.K., with this increase disproportionately affecting London. A recent national survey suggests that co-infection with HIV occurs in less than 5% of tuberculosis patients. This study asked if local co-infection rates in Inner London differed from the national results. METHODS: 157 consecutive patients starting antituberculous chemotherapy were venesected 2 weeks into treatment. Anonymized blood samples were screened for antibodies for HIV-1 and HIV-2 by enzyme-linked immunosorbent assay (ELISA). Epidemiological data were collected on each patient which was also coded before HIV test results were known. RESULTS: Of 157 patients commencing antituberculous therapy, 39 patients (24.8%) were found to be co-infected with HIV-1. HIV-negative and positive patients were similar in terms of age and sex. When 98 patients giving their country of origin as other than Europe were considered there were 22 co-infected with HIV (22.4%). Of the 39 HIV-positive identified in this study, 37 were also identified by our voluntary HIV testing programme. CONCLUSIONS: This study has shown that there may be very different rates of co-infection at a local level in the U.K. The local variation may be missed by national surveys and diverse local testing procedures. Anonymous testing identified only two patients with tuberculosis and HIV infection who were not identified by our voluntary HIV testing programme and this suggests that offering HIV tests to patients with tuberculosis is largely taken up by those at risk of HIV infection. Surveillance studies of this type are important in identifying marked local variation from the national pattern of HIV and Mycobacterium tuberculosis infection.

Pulmonary tuberculosis in Harare, Zimbabwe: analysis by spoligotyping.

BACKGROUND: Over the last 10 years there has been a fourfold increase in cases of tuberculosis in Harare, Zimbabwe. The use of molecular epidemiology to understand tuberculosis transmission in this epidemic has been hampered by the availability of suitable culture facilities. A study was therefore undertaken to explore the potential of spoligotyping, a polymerase chain reaction based technique that does not require tuberculosis culture. METHODS: Adults attending a chest clinic with clinical or radiological pulmonary tuberculosis and one smear positive sputum were enrolled over one month. Demographic, socioeconomic, and clinical data were gathered using a standardised questionnaire. Molecular fingerprinting of genomic DNA recovered from sputum was performed by spoligotyping. RESULTS: Sixty one subjects (median age 28 years (range 18-73); 61% men) were recruited and 57 provided adequate sputum samples. Recent rural-urban migration or immigration was not common; 40% of subjects lived in crowded living conditions. DNA suitable for spoligotyping was recovered from 28 patients and 20 different genotypes of Mycobacterium tuberculosis were identified. Fifteen patients were infected with an M tuberculosis strain shared by one or more individuals. Patients infected with a shared spoligotype were not closely linked geographically within Harare, but were more likely to live in overcrowded conditions (69% versus 23%; odds ratio 6.85 (95% CI 1.2 to 47), p = 0.026). Analysis of the patients' original rural family homes revealed two geographically related spoligotype clusters. CONCLUSIONS: Spoligotyping may yield valuable molecular typing information in populations where tuberculosis culture is not available. This novel technique requires further development and evaluation in larger epidemiological studies.

Production of tumor necrosis factor and nitric oxide by macrophages infected with live and dead mycobacteria and their suppression by an interleukin-10-secreting recombinant.

We have analyzed mycobacterium-induced cytokine secretion in the J774A.1 macrophage-like cell line. Tumor necrosis factor alpha (TNF-alpha) was preferentially induced by live organisms, both slow and rapid growing. Expression of interleukin-10 by a recombinant strain of Mycobacterium smegmatis caused reduced production of TNF-alpha and nitric oxide during the early stages of infection.

Differential responses to challenge with live and dead Mycobacterium bovis Bacillus Calmette-Guérin.

Bacillus Calmette-Guérin (BCG) vaccination has been shown to protect against challenge with virulent Mycobacterium tuberculosis in a range of experimental animal models: in each case, protective efficacy requires vaccination with live bacteria. With the goal of moving to a new generation of safer, nonliving vaccines, efforts have been made to identify the factors that determine the efficacy of live vaccination. We show that injection of live, but not dead, BCG induces localized swelling in the mouse footpad model. Live and dead bacteria induce similar responses during the first week after vaccination as determined by immunohistochemical analysis of the site of injection and of the draining lymph node. The subsequent differential response is characterized by migration of acid-fast bacilli to the draining lymph node in the case of the live vaccine. This is accompanied by an increase in mononuclear cells in the lymph node and by expression of inducible nitric oxide synthase (iNOS) both in the lymph node and at the site of injection. The ability of the bacteria to migrate to the lymph node may be an important element in the efficacy of live BCG vaccination.

Increased inflammatory cytokines and new collagen formation in cutaneous tuberculosis and sarcoidosis.

BACKGROUND: Interactions between mononuclear cells, vascular endothelium, fibroblasts, and cytokines during the inflammatory reaction within a granuloma have the potential to contribute to the progression to fibrosis. METHODS: Biopsy specimens of six tuberculous and eight sarcoidosis skin lesions were examined by immunohistochemistry to seek evidence for the presence of inflammatory and fibrotic reactions in human granulomatous disease. Additionally, to understand how a T cell mediated delayed type hypersensitivity reaction--a component of chronic granulomatous inflammation--could progress to fibrosis, the human in vivo model of the cutaneous tuberculin Heaf reaction to purified protein derivative (PPD) was studied in a group of 48 subjects. RESULTS: Granulomas from tuberculous and sarcoidosis skin biopsy specimens were seen to contain cells with marked staining by antibodies to fibronectin, transforming growth factor beta (pan TGF-beta), and type 1 procollagen (PCP-1). Accentuated staining of extracellular matrix was seen both in the granulomas and in the peri-granulomatous regions. Less prominent staining was observed using antibodies against interleukin 1 beta (IL-1 beta) and alpha-smooth muscle actin (alpha-SMA). Biopsies of Heaf reactions revealed cells staining for IL-1 beta, tumour necrosis factor alpha (TNF-alpha), platelet derived growth factor B (PDGF-B), and fibronectin which were detected as early as day 1 and persisted throughout the 14 day study period. Cells staining for PCP-1 increased to greatest abundance at day 14. All these cytokines were present in low abundance in biopsy specimens from sites inoculated with saline only. CONCLUSIONS: Evidence is provided that granulomas in tuberculosis and sarcoidosis behave as active centres of fibrogenesis. Using the Heaf model, the temporal relationship between the early appearance of cytokines and the later increase in the collagen precursor PCP-1 linked the immune mediated chronic inflammatory response with subsequent fibrosis and suggested that the tuberculin Heaf reaction will serve as a model for studying the early events of granuloma formation in patients with tuberculosis and sarcoidosis.