Gr1int/high Cells Dominate the Early Phagocyte Response to Mycobacterial Lung Infection in Mice.

Lung infection by Mycobacterium tuberculosis is characterized by chronic infection of lung-resident macrophages, long considered to be the primary hosts and determinants of the outcome of the early immune response. Although alveolar macrophages are well-known to host intracellular mycobacteria at later stages of disease, little is known about the earliest events of the innate immune response. The phagocytes that take up mycobacteria immediately following infection, and how the early lung phagocyte response is altered by vaccination with M. bovis bacille Calmette-Guérin (BCG) were unknown. Using BCG expressing the bright red fluorescent protein tdTomato and flow cytometry, we modeled early infection in C57BL/6 mice and tracked phagocyte population kinetics and uptake of mycobacteria, to better understand the involvement of specific phagocyte subsets. By 1 day post-infection, dose-dependent accumulation of neutrophils was observed and surprisingly, granulocytes comprised a greater proportion of infected phagocytes than alveolar macrophages. By 7 days post-infection alveolar macrophages had become the dominant BCG-associated phagocytes. Prior mucosal BCG exposure provided immunized mice with greater frequencies and numbers of lung macrophage subsets, and a significantly greater proportion of alveolar macrophages expressed CD11b prior to and following challenge infection. These data provide the first evidence of granulocytes as the dominant infected phagocyte subset early after mycobacterial infection, and highlight enhanced recruitment of lung macrophages as a factor associated with protection in BCG-immunized mice.

Do surgical helmet systems affect intraoperative wound contamination? A randomised controlled trial.

BACKGROUND: Deep infection following arthroplasty remains a devastating complication. Some registry data suggests that modern positive-pressure surgical helmet systems (SHS) are associated with a paradoxical increase in infection rates, and as such their role in arthroplasty remains unclear. The aim of this study was to investigate whether SHS increase wound contamination in total knee arthroplasty (TKA) and if this contamination can be reduced by placing tape around the gown/glove interface. METHODS: Seventy-five patients were randomised into three groups: scrubbed theatre staff wore standard surgical gowns (SG), SHS without tape at the gown/glove interface, or SHS with tape. All TKA operations were carried out by the same surgeon. Wound contamination was assessed using a wound culture technique. Blinded laboratory analysis was performed. RESULTS: There were 5/50 culture positive cases when a SHS was used compared to 0/25 when a SG was used; but this difference was not statistically significant (p = 0.16). There were 4/24 culture positive cases when SHS with tape was used compared to 1/26 when SHS without tape was used; but this difference was not statistical significant p = 0.18. CONCLUSION: We found no difference in wound contamination between SG and SHS. Addition of tape at the gown/glove interface did not alter the contamination rate. The choice of surgical gown should take into account cost, comfort and personal protection; as this study found no evidence that wound contamination rates will be altered.

A Novel Restraint Device for Injection of Galleria mellonella Larvae that Minimizes the Risk of Accidental Operator Needle Stick Injury.

Larvae of the insect Galleria mellonella are increasingly being used for studying pathogenic microbes and their virulence mechanisms, and as a rapid model for screening novel antimicrobial agents. The larvae (waxworms) are most frequently infected by injection of pathogenic organisms into the haemocoel through the insect's prolegs. The mostly widely used method for restraining the waxworms for injection is by grasping them between the operator's fingers, which puts the operator at risk of needle stick injury, an important consideration when working with highly pathogenic and/or drug-resistant microorganisms. While use of a stab proof glove can reduce this risk of injury, it does so at the loss of manual dexterity and speed, resulting in a more labor-intensive, and cumbersome assay. We describe a simple cost effective device (the so-called "Galleria Grabber") for restraining waxworms for injection that keeps the operator's fingers clear of the needle thus reducing the risk of injury.

Effect of common and experimental anti-tuberculosis treatments on Mycobacterium tuberculosis growing as biofilms.

Much is known regarding the antibiotic susceptibility of planktonic cultures of Mycobacterium tuberculosis, the bacterium responsible for the lung disease tuberculosis (TB). As planktonically-grown M. tuberculosis are unlikely to be entirely representative of the bacterium during infection, we set out to determine how effective a range of anti-mycobacterial treatments were against M. tuberculosis growing as a biofilm, a bacterial phenotype known to be more resistant to antibiotic treatment. Light levels from bioluminescently-labelled M. tuberculosis H37Rv (strain BSG001) were used as a surrogate for bacterial viability, and were monitored before and after one week of treatment. After treatment, biofilms were disrupted, washed and inoculated into fresh broth and plated onto solid media to rescue any surviving bacteria. We found that in this phenotypic state M. tuberculosis was resistant to the majority of the compounds tested. Minimum inhibitory concentrations (MICs) increased by 20-fold to greater than 1,000-fold, underlying the potential of this phenotype to cause significant problems during treatment.

Biologically Active Acetylenic Amino Alcohol and N-Hydroxylated 1,2,3,4-Tetrahydro-ß-carboline Constituents of the New Zealand Ascidian Pseudodistoma opacum.

The first occurrence of an acetylenic 1-amino-2-alcohol, distaminolyne A (1), isolated from the New Zealand ascidian Pseudodistoma opacum, is reported. The isolation and structure elucidation of 1 and assignment of absolute configuration using the exciton coupled circular dichroism technique are described. In addition, a new N-9 hydroxy analogue (2) of the known P. opacum metabolite 7-bromohomotrypargine is also reported. Antimicrobial screening identified modest activity of 1 toward Escherichia coli, Staphylococcus aureus, and Mycobacterim tuberculosis, while 2 exhibited a moderate antimalarial activity (IC50 3.82 µM) toward a chloroquine-resistant strain (FcB1) of Plasmodium falciparum.

Regional Intraosseous Administration of Prophylactic Antibiotics is More Effective Than Systemic Administration in a Mouse Model of TKA.

BACKGROUND: In human TKA studies, intraosseous regional administration (IORA) of prophylactic antibiotics achieves local tissue antibiotic concentrations 10 times greater than systemic administration. However, it is unclear if such high concentrations provide more effective prophylaxis. QUESTIONS/PURPOSES: We asked: (1) What prophylaxis dosage and route (intravenous [IV] versus IORA of prophylactic antibiotics) produce less in vivo bacterial burden compared with no-antibiotic controls? (2) Compared with controls, what prophylaxis dosage and route yield fewer colony-forming units (CFUs) in euthanized animals in a model of TKA? (3) Is prophylactic IORA of antibiotics more effective than same-dose IV antibiotic administration in reducing CFUs? METHODS: Mice (six to nine per group) were block randomized to one of six prophylaxis regimens: control, systemic cefazolin (C100IV), IORA of cefazolin (C100IORA), systemic vancomycin (V110IV), low-dose systemic vancomycin (V25IV), and low-dose IORA of vancomycin (V25IORA). Surgery involved placement of an intraarticular knee prosthesis, followed by an inoculum of bioluminescent Staphylococcus aureus strain Xen36. Biophotonic imaging assessed in vivo bacterial loads, and after 4 days bacterial load was quantified using culture-based techniques. Comparisons were made for each prophylactic regimen to controls and between same-dose IV and IORA of prophylactic antibiotic regimens. RESULTS: Mice treated with systemic high-dose vancomycin, IORA of vancomycin, or IORA of cefazolin had lower in vivo Staphylococcus aureus burdens (median area under curve, Control: 5.0 × 10(6); V110IV: 1.5 × 10(6), difference of medians 3.5 × 10(6), p = 0.003; V25IV: 1.94 × 10(6), difference 3.07 × 10(6), p = 0.49; V25IORA: 1.51 × 10(6), difference 3.5 × 10(6), p = 0.0011; C100IORA: 1.55 × 10(6), difference 3.46 × 10(6), p = 0.0016; C100IV: 2.35 × 10(6), difference 2.66 × 10(6), p = 0.23.) Similar findings were seen with culture-based techniques on recovered implants. IORA of prophylactic antibiotics was more effective than same-dose IV administration in reducing bacterial load on recovered implants (median CFUs < 7.0 × 10(0) vs 2.83 × 10(2), p = 0.0183). CONCLUSIONS: IORA of prophylactic cefazolin and vancomycin was more effective than the same dose of antibiotic given systemically. The effectiveness of vancomycin in particular was enhanced by IORA of prophylactic antibiotics despite using a lower dose. CLINICAL RELEVANCE: Our study supports previous studies of IORA of prophylactic antibiotics in humans and suggests this novel form of administration has the potential to enhance the effectiveness of prophylaxis in TKA. Because of concerns regarding antibiotic stewardship, IORA of prophylactic vancomycin may be more appropriately restricted to patients having TKA who are at greater risk of infection, and clinical trials are in progress.

Detection of Mycobacterium avium subspecies paratuberculosis in patients with Crohn's disease is unrelated to the presence of single nucleotide polymorphisms rs2241880 (ATG16L1) and rs10045431 (IL12B).

Mycobacterium avium subspecies paratuberculosis (MAP) has been controversially linked with Crohn's disease (CD). Detection of MAP in CD has been highly variable, and one explanation might be the genetic heterogeneity of this syndrome. Many of the single nucleotide polymorphisms (SNPs) linked with CD are contained within genes that are associated with bacterial handling in general, and some are specifically implicated in susceptibility to mycobacterial disease. We tested a cohort of IBD patients (n = 149) to determine whether the presence of MAP was associated with a selection of these SNPs. Blood samples from CD patients (n = 84), ulcerative colitis (UC, n = 65) patients and healthy controls (n = 55) were examined for the presence of MAP and SNPs in ATG16L1, IL12B, NOD2/CARD15, NKx2-3, IL23R and IRGM. Statistical analysis was then used to determine whether there was any association between the presence of MAP and these SNPs. MAP, rs2241880 (ATG16L1) and rs10045431 (IL12B) were found to be significantly associated with CD. The presence of MAP was not related to the status of the SNPs in ATG16L1 or IL12B. We have found no evidence for the contribution of these SNPs to the presence of MAP in CD patients.