Genetic and biochemical characterization of BIM-1, a novel acquired subgroup B1 MBL found in a Pseudomonas sp. strain from the Brazilian Amazon region.

OBJECTIVES: To characterize a novel acquired MBL, BIM-1, in a Pseudomonas #2 (subgroup P. guariconensis) strain isolated from the Aurá river located in the Brazilian Amazon hydrographic basin. METHODS: WGS using an Illumina® MiSeq System was used to characterize the genome of Pseudomonas sp. IEC33019 strain. Southern blotting/hybridization assays were performed to confirm the location of the MBL-encoding gene, blaBIM-1 (Belém Imipenemase). Antimicrobial susceptibility testing, cloning, and biochemical and phenotypic characterization were performed to determine BIM-1 kinetics. RESULTS: The IEC33019 strain showed high resistance rates to ß-lactams, ciprofloxacin and aminoglycosides, being susceptible only to polymyxins and susceptible, increased exposure to aztreonam. WGS analysis revealed a novel acquired MBL-encoding gene, blaBIM-1, found as a gene cassette inserted into a class 1 integron (In1326) that also carried qnrVC1 and aadA11e. In1326 was located in a complex transposon, Tn7122, carried by a 52.7 kb conjugative plasmid (pIEC33019) with a toxin/antitoxin system (vapB/vapC). BIM-1 belongs to the molecular subgroup B1 and shares 70.2% and 64.9% similarity with SIM-1 and IMP-1, respectively. Kinetics analysis of BIM-1 showed hydrolytic activity against all ß-lactams tested. CONCLUSIONS: BIM-1 is a novel acquired MBL encoded by a gene carried by mobile genetic elements, which can be transferred to other Gram-negative bacilli (GNB). Because the IEC33019 strain was recovered from a river impacted by a populous metropolitan region with poor basic sanitation and served by limited potable freshwater, it would be important to establish the role of the BIM-1-producing GNB as nosocomial pathogens and/or as colonizers of the riverside population in this geographical region.

Streptococcus pneumoniae serotype 19A from carriers and invasive disease: virulence gene profile and pathogenicity in a Galleria mellonella model.

PURPOSE: This study aimed to evaluate and compare the presence of genes related to surface proteins between isolates of Streptococcus pneumoniae from healthy carriers (HC) and invasive pneumococcal disease (IPD) with a particular focus on serotype 19A. METHODS: The presence of these genes was identified by real-time PCR. Subsequently, we employed the Galleria mellonella larval infection model to study their effect on pathogenicity in vivo. RESULTS: The percentage of selected virulence genes was similar between the HC and IPD groups (p > 0.05), and the genes lytA, nanB, pavA, pcpA, phtA, phtB, phtE, rrgA, and sipA were all present in both groups. However, the virulence profile of the isolates differed individually between HC and IPD groups. The highest lethality in G. mellonella was for IPD isolates (p < 0.01), even when the virulence profile was the same as compared to the HC isolates or when the nanA, pspA, pspA-fam1, and pspC genes were not present. CONCLUSIONS: The occurrence of the investigated virulence genes was similar between HC and IPD S. pneumoniae serotype 19A groups. However, the IPD isolates showed a higher lethality in the alternative G. mellonella model than the HC isolates, regardless of the virulence gene composition, indicating that other virulence factors may play a decisive role in virulence. Currently, this is the first report using the in vivo G. mellonella model to study the virulence of clinical isolates of S. pneumoniae.

Adaptive HIV-specific B cell-derived humoral immune defenses of the intestinal mucosa in children exposed to HIV via breast-feeding.

BACKGROUND: We evaluated whether B cell-derived immune defenses of the gastro-intestinal tract are activated to produce HIV-specific antibodies in children continuously exposed to HIV via breast-feeding. METHODS: Couples of HIV-1-infected mothers (n = 14) and their breastfed non HIV-infected (n = 8) and HIV-infected (n = 6) babies, and healthy HIV-negative mothers and breastfed babies (n = 10) as controls, were prospectively included at the Complexe Pédiatrique of Bangui, Central African Republic. Immunoglobulins (IgA, IgG and IgM) and anti-gp160 antibodies from mother's milk and stools of breastfed children were quantified by ELISA. Immunoaffinity purified anti-gp160 antibodies were characterized functionally regarding their capacity to reduce attachment and/or infection of R5- and X4- tropic HIV-1 strains on human colorectal epithelial HT29 cells line or monocyte-derived-macrophages (MDM). RESULTS: The levels of total IgA and IgG were increased in milk of HIV-infected mothers and stools of HIV-exposed children, indicating the activation of B cell-derived mucosal immunity. Breast milk samples as well as stool samples from HIV-negative and HIV-infected babies exposed to HIV by breast-feeding, contained high levels of HIV-specific antibodies, mainly IgG antibodies, less frequently IgA antibodies, and rarely IgM antibodies. Relative ratios of excretion by reference to lactoferrin calculated for HIV-specific IgA, IgG and IgM in stools of HIV-exposed children were largely superior to 1, indicating active production of HIV-specific antibodies by the intestinal mucosa. Antibodies to gp160 purified from pooled stools of HIV-exposed breastfed children inhibited the attachment of HIV-1NDK on HT29 cells by 63% and on MDM by 77%, and the attachment of HIV-1JRCSF on MDM by 40%; and the infection of MDM by HIV-1JRCSF by 93%. CONCLUSIONS: The intestinal mucosa of children exposed to HIV by breast-feeding produces HIV-specific antibodies harbouring in vitro major functional properties against HIV. These observations lay the conceptual basis for the design of a prophylactic vaccine against HIV in exposed children.

The prevalence of hepatitis B virus markers in a cohort of students in Bangui, Central African Republic.

BACKGROUND: Hepatitis B virus (HBV) is the major cause of chronic hepatitis, cirrhosis, and hepatocellular carcinoma. The global epidemiological scenario of HBV infection has been changing rapidly over the last two decades due to an effective immunization programme initiated by the World Health Organization. The objective of this study is to estimate the prevalence of HBV in apparently healthy young people and to identify the risk factors of transmission of the HBV among this population in Bangui. METHODS: Dried blood Spots from 801 adolescent high school and young adult university students were prepared by spotting a drop of whole blood (4 spots) from the same fingerprick onto Whatman filter paper. A blood sample aliquot eluted from DBS was then processed with commercial ELISA tests (Abbott Murex, Dartfort, UK) to detect HBsAg antigen, Anti-HBc and Anti-HBs antibodies). RESULTS: The overall prevalence was 42.3% for antibody to hepatitis B core antigen, 15.5% for HBsAg of which 1.3% of HBsAg alone. HBV familial antecedents, sexual activity and socioeconomic conditions were the main risk factors of HBV infection encountered in the adolescents and young adults. CONCLUSION: These results show for the first time the high prevalence of HBV in apparently healthy young people in Bangui. This high prevalence is age- and sex-independent. Transmission risk factors were a familial antecedent of HBV, no utilisation of condoms and public scholarship. To lower HBV prevalence, an adequate program of active screening and vaccination for adolescents and young adults should be implemented, along with a universal immunization program.

Broad spectrum of coreceptor usage and rapid disease progression in HIV-1-infected individuals from Central African Republic.

To study the progression of HIV-1 infection and coreceptor usages in Central African Republic, clinical data, plasma viral load, and coreceptor usage of sequential HIV-1 isolates were analyzed in a seroincident prospective cohort (PRIMOCA). Twenty-three HIV-1 infected individuals from the Central African Armed Forces were followed from 1995 to 2000. Viruses were isolated from 17 patients at various time points after seroconversion and their coreceptor usage was examined using GHOST cells expressing CD4 and one of the HIV-1 chemokine coreceptors CCR5, CXCR4, BOB/GPR15, and Bonzo/STRL33/CXCR6. Eleven patients died from AIDS. Eight of them died between 2 and 5 years after seroconversion, after a brief symptomatic stage. Patients who rapidly progressed to AIDS and death displayed the highest viral loads after seroconversion. All isolates obtained soon after seroconversion used CCR5, albeit, in some cases, CXCR4, BOB, or Bonzo were also used. Most isolates remained R5 (59 out of 61 isolates), although viruses using CXCR4 appeared in some cases of progression to AIDS. In several cases, a broad tropism was observed during the course of infection, with a frequent usage of BOB and Bonzo in addition to CCR5. Rapid progression to disease and short survival time among Central African HIV-1 patients appear more frequent than those reported in industrialized countries. Viral coreceptor used was mainly CCR5, but, interestingly, a large part of isolates also used BOB and Bonzo. However, there was no strict correlation between the clinical outcome and extended viral tropism.