Seroprevalence and molecular characterization of viral hepatitis and HIV co-infection in the Central African Republic.

BACKGROUND: The Central African Republic (CAR) is one of the countries with the highest prevalence of viral hepatitis infection in the world. Coinfection with HIV increases the morbidity and mortality beyond that of mono-infection with either hepatitis or HIV. The present study describes the geographic distribution of viral hepatitis infections and molecular characterization of these viruses in the CAR. METHODOLOGY: Out of 12,599 persons enrolled during the fourth Multiple Indicator Cluster Survey of 2010 in the CAR, 10,621 Dried Blood Spot (DBS) samples were obtained and stored at -20°C. Of these DBS, 4,317 samples were randomly selected to represent all regions of the CAR. Serological tests for hepatitis B, D, and C viruses were performed using the ELISA technique. Molecular characterization was performed to identify strains. RESULTS: Of the 4,317 samples included, 53.2% were from men and 46.8% from women. The HBsAg prevalence among participants was 12.9% and that HBc-Ab was 19.7%. The overall prevalence of HCV was 0.6%. Co-infection of HIV/HBV was 1.1% and that of HBV/HDV was 16.6%. A total of 77 HBV, 6 HIV, and 6 HDV strains were successfully sequenced, with 72 HBV (93.5%) strains belonging to genotype E and 5 (6.5%) strains belonging to genotype D. The 6 HDV strains all belonged to clade 1, while 4 recombinants subtype were identified among the 6 strains of HIV. CONCLUSION: Our study found a high prevalence of HBV, HBV/HDV and HBV/HIV co-infection, but a low prevalence of HCV. CAR remains an area of high HBV endemicity. This study's data and analyses would be useful for establishing an integrated viral hepatitis and HIV surveillance program in the CAR.

Colistin Resistance Mechanism in Enterobacter hormaechei subsp. steigerwaltii Isolated from Wild Boar (Sus scrofa) in France.

Wild animals may act as efficient antimicrobial-resistance reservoirs and epidemiological links between humans, livestock, and natural environments. By using phenotypic and genotypic characterization, the present study highlighted the occurrence of an antimicrobial-resistant (i.e., amoxicillin, amoxicillin-clavulanic acid, cephalothin, and colistin) Enterobacter hormaechei subsp. steigerwaltii strain in wild boar (Sus scrofa) from France. The molecular analysis conducted showed non-synonymous mutations in the pmrA/pmrB and phoQ/phoP operons and the phoP/Q regulator mgrB gene, leading to colistin resistance. The present data highlight the need for continuous monitoring of multidrug-resistant bacteria in wild animals to limit the spread of these threatening pathogens.

Molecular Characterization of Multidrug-Resistant Escherichia coli Isolated from Milk of Dairy Cows with Clinical Mastitis in Algeria.

The objective of this study was to investigate the occurrence of multidrug-resistant Escherichia coli in cows with clinical mastitis in 42 different dairy farms located in the Bordj Bou Arreridj region of Algeria. Milk samples were cultured on Columbia blood agar, and isolates were then identified by matrix-assisted laser desorption ionization-time of flight mass spectrometry. In total, 200 samples were screened and 52 E. coli strains confirmed as causative agents were obtained. The antimicrobial susceptibility testing was performed by disk diffusion method. Antibiotic resistance genes, including those conferring resistance to extended-spectrum ß-lactamases (i.e., blaTEM, blaSHV, and blaCTX-M), tetracyclines (tetA, tetB, tetC, and tetJ), aminoglycosides [aph(3'), aac(3'), aac(6'), ant, aad, and armA], and quinolones (qnrA and qnrB) were amplified by standard PCR and sequenced when positive. Transferability of resistance genes has been investigated by conjugation experiments and multilocus sequence typing. The most frequently observed resistance was to amoxicillin (86.5%), followed by tetracycline (75%), amoxicillin-clavulanic acid (59.6%), trimethoprim-sulfamethoxazole (36.5%), doxycycline (13.5%), and ciprofloxacin (13.5%). Multidrug resistance was observed in 38.4% of isolates. Genotypic characterization showed that tetA (44.2%) and blaTEM-1 (30.7%) genes were the most prevalent. Screening for plasmid-mediated quinolone resistance genes demonstrated that seven isolates (13.5%) expressed qnrB and one isolate (1.9%) harbored qnrA. In addition, aminoglycoside resistance determinants including aadA1 and aac(3)-Id were detected in seven and two isolates, respectively. Moreover, blaTEM, tetA, tetB, qnrB, and aadA1 were successfully transferred horizontally to transconjugant strains. The multilocus sequence typing revealed the presence of three different sequence types (ST162, ST371, and ST 949).

Development of real-time PCR assay allowed describing the first clinical Klebsiella pneumoniae isolate harboring plasmid-mediated colistin resistance mcr-8 gene in Algeria.

OBJECTIVES: We aimed to develop here a specific real-time PCR assay with TaqMan® probe to detect efficiently bacterial strains harboring the new plasmid mediated-colistin resistance mcr-8 gene. METHODS: Specific primers and probe for mcr-8 gene were designed from sequences alignment of all mcr genes variants. Specificity of the designed primers and probe were first checked par BlastN analysis and by in silico PCR. The analytical sensitivity and specificity tests were performed in vitro on a panel of 290 genomic DNA of Gram-negative bacteria and 250 metagenomic DNA from human stool samples. Whole genome sequencing (WGS) was performed here using MiSeq technology. RESULTS: Designed primers and probe were 100% specific tomcr-8 gene by BlastN and in silico PCR analysis. Real-time PCR screening of a collection of clinical isolates resulted to one positive Klebsiella pneumoniae isolate (KP95). WGS confirmed that this isolate harbored the mcr-8 gene and other resistance genes such as blaOXA-48, blaCTX-M-15 ß-lactamases. Our real-time PCR was highly sensitive on a 10-fold dilution serie from a calibrated inoculum at 108 CFU/mL with a limit of detection at 55 CFU/mL. CONCLUSION: To the best of our knowledge, we propose here, the first real-time PCR assay targeting mcr-8 gene with high specificity and sensitivity, able to detect mcr-8 gene in less than 2 h from any DNA sample. This real-time PCR assay allowed the first description of a clinical K. pneumoniae strain harboring the mcr-8 gene in Algeria.

Investigation of urban birds as source of ß-lactamase-producing Gram-negative bacteria in Marseille city, France.

BACKGROUND: We investigate here the presence of multidrug-resistant bacteria isolated from stool samples of yellow-legged gulls and chickens (n = 136) in urban parks and beaches of Marseille, France. Bacterial isolation was performed on selective media, including MacConkey agar with ceftriaxone and LBJMR medium. Antibiotic resistance genes, including extended-spectrum ß-lactamases (ESBL) (i.e. blaCTX-M, blaTEM and blaSHV), carbapenemases (blaKPC, blaVIM, blaNDM, blaOXA-23, blaOXA-24, blaOXA-48 and blaOXA-58) and colistin resistance genes (mcr-1 to mcr-5) were screened by real-time PCR and standard PCR and sequenced when found. RESULTS: Of the 136 stools samples collected, seven ESBL-producing Gram-negative bacteria (BGN) and 12 colistin-resistant Enterobacteriaceae were isolated. Among them, five ESBL-producing Escherichia coli and eight colistin-resistant Hafnia alvei strains were identified. Four blaTEM-1 genes were detected in yellow-legged gulls and chickens. Three CTX-M-15 genes were detected in yellow-legged gulls and pigeons, and one CTX-M-1 in a yellow-legged gull. No mcr-1 to mcr-5 gene were detected in colistin-resistant isolates. Genotyping of E. coli strains revealed four different sequence types already described in humans and animals and one new sequence type. CONCLUSIONS: Urban birds, which are believed to have no contact with antibiotics appear as potential source of ESBL genes. Our findings highlight the important role of urban birds in the proliferation of multidrug-resistant bacteria and also the possible zoonotic transmission of such bacteria from wild birds to humans.