Short-term effects of amoxicillin on bacterial communities in manured soil.

Antibiotic-resistant bacteria, nutrients and antibiotics that enter the soil by means of manure may enhance the proportion of bacteria displaying antibiotic resistance among soil bacteria and may affect bacterial community structure and function. To investigate the effect of manure and amoxicillin added to manure on soil bacterial communities, microcosm experiments were performed with two soil types and the following treatments: (1) nontreated, (2) manure-treated, (3) treated with manure supplemented with 10 mg amoxicillin kg(-1) soil and (4) treated with manure supplemented with 100 mg amoxicillin kg(-1) soil, with four replicates per treatment. Manure significantly increased the total CFU count and the amoxicillin-resistant CFU count of both soil types. However, only the soil with a history of manure treatment showed a significant increase in the relative number of amoxicillin-resistant bacteria as a result of amoxicillin amendment. The majority of plasmids exogenously isolated from soil originated from soil treated with amoxicillin-supplemented manure. All 16 characterized plasmids carried the bla-TEM gene, and 10 of them belonged to the IncN group. The bla-TEM gene was detected in DNA directly extracted from soil by dot-blot hybridization of PCR amplicons and showed an increased abundance in soil samples treated with manure. Molecular fingerprint analysis of 16S rRNA gene fragments amplified from soil DNA revealed significant effects of manure and amoxicillin on the bacterial community of both soils.

Clonally expanded CD4+CD28null T cells in rheumatoid arthritis use distinct combinations of T cell receptor BV and BJ elements.

Clonally expanded, autoreactive CD4(+)CD28(null) cells can be found in the peripheral blood of patients with rheumatoid arthritis and have been shown to be associated with severeextra-articular disease manifestations. We investigated the size of the CD4(+)CD28(null) compartment and the TCR beta chain repertoire of expanded CD4(+) clonotypes in 94 rheumatoid arthritis patients by complementarity-determining region 3 (CDR3) length analysis (spectratyping) in the BV6 and BV14 TCR families, with primers specific for three arbitrarily chosen beta chain joining elements (BJ1S2, BJ2S3 and BJ2S7). The spectratyping results showed a strong correlation of the size of the CD4(+)CD28(null) compartment with the detected number of BV14 clonotypes, whereas no association with BV6 oligoclonality was found. Only clones using the BV14-BJ1S2 and BV14-BJ2S3 combinations contributed to this correlation, however, whereas BV14-BJ2S7 clones did not. This preferential correlation implies a role for the TCR beta chain in stimulating clonal outgrowth and argues against the previously suggested superantigenic stimulation of in-vivo-expanded clones. Instead, since no evidence for shared antigen specificity could be detected, clonal expansion of T cells in rheumatoid arthritis might be influenced by the BJ elements because of changes in the flexibility of the protein backbone of the beta-chain.

B lymphocytopenia in rheumatoid arthritis is associated with the DRB1 shared epitope and increased acute phase response.

The influence of HLA DRB1 alleles on B-cell homeostasis was analyzed in 164 patients with rheumatoid arthritis (RA). The percentages of CD19+ B lymphocytes determined in the peripheral circulation of 94 retrospectively recruited RA patients followed a bimodal distribution. Two frequency peaks (B-cell(low) patients and B-cell(high) patients) were separated by the population median of a B-cell frequency of 8.5% of all lymphocytes. Human leucocyte antigen genotyping revealed that the B-cell(low) patients were more frequently positive for the RA-associated HLA DRB1 shared epitope (SE) than were B-cell(high) patients. Accordingly, SE-positive patients had lower CD19 percentages in the rank-sum analysis when compared with SE-negative patients, and were markedly B lymphocytopenic when compared with a healthy control group. To confirm the differential frequencies of CD19+ B cells, absolute numbers in peripheral blood were determined prospectively in a cohort of 70 RA patients with recent onset disease. SE-positive patients were found to have lower absolute numbers of circulating CD19+ B cells. B-cell counts below the mean of the study population were associated with higher acute phase response and with increased levels of rheumatoid factor IgA. No correlation between absolute numbers of circulating B cells and radiographic progression of joint destruction was seen. The influence of immunogenetic parameters on B-cell homeostasis in RA reported here has not been described previously. The clinical relevance of B lymphocytopenia in SE-positive RA will be further investigated in longitudinal studies.