- Volume 62, Issue 11, 2013

The natural habitat of the multiflagellate protozoon Lophomonas blattarum is as an endocommensal in the hindgut of insects such as cockroaches. However, it also causes bronchopulmonary disease in humans. The aim of this paper was to review the literature on this organism in the context of respiratory disease. The biology epidemiology, route of transmission, pathogenic mechanisms and diagnosis methods are also described. A total of 61 cases were identified in the literature. The majority of these reports were from China, with some cases from Peru and Spain. Most cases were adult males, although paediatric cases were reported in Peru. Clinical presentation was non-specific, including symptoms such as fever, cough and breathless. Antiprotozoal therapy was generally effective.

Mortality from bloodstream infections (BSIs) correlates with diagnostic delay and the use of inappropriate empirical treatment. Early PCR-based diagnosis could decrease inappropriate treatment, improving patient outcome. The aim of the present study was to assess the clinical utility of this molecular technology to diagnose BSIs. We assessed a new dual-priming oligonucleotide-based multiplex PCR assay, the Magicplex Sepsis Test (MST) (Seegene), along with blood culture (BC). A total of 267 patients from the intensive care unit and haematology and emergency departments were enrolled. Clinical data were also used by physicians to determine the likelihood of infection. Ninety-eight (37 %) specimens were positive: 29 (11 %) by both the MST and BC, 29 (11 %) by the MST only, and 40 (15 %) by BC only. The proportion of agreement between the two methods was 73 % (Cohen’s κ: 0.45; 0.28–0.6; indicating fair to moderate agreement). According to clinical assessment, 63 (64 %) positive specimens were considered BSIs: 23 (36 %) were positive by both the MST and BC, 22 (35 %) were positive only by BC, and 18 (29 %) were positive only by the MST. Thirty-eight (14 %) positive specimens by the MST and/or BC were considered as contaminants. Of 101 specimens collected from patients receiving antibiotics, 20 (20 %) were positive by the MST and 32 (32 %) by BC. Sensitivity and specificity were 65 % and 92 %, respectively, for the MST and 71 % and 88 %, respectively for BC. We concluded that the MST shows a high specificity but changes in design are needed to increase bacteraemia detection. For viability in clinical laboratories, technical improvements are also required to further automate the process.

Human papillomavirus type 16 (HPV16) non-European variants have been associated with persistent infection and cervical cancer development, while the L83V variant of the E6 gene has been correlated with the progression of cervical malignancy. The present study investigated the presence of the HPV16 L83V variant in Greek women. Molecular evolutionary analysis of the HPV16 E6 and E7 oncogenes was conducted in order to estimate the evolution of the HPV16 genome in the Greek population. The E6 L83V variant was found in 78.2 % of high- and 64.28 % of low-grade specimens. Moreover, the prototype and E6 L83V variants were both prevalent in high- and low-grade malignancies in Greek women. Selective pressure analysis of the individual amino acid residues of HPV16 sequences from the Greek population indicates that codon 83 of the E6 protein, as well as codon 85 of the E7 protein, are undergoing positive selection. Novel sequence variations were recorded within the E6 and E7 genes in cervical samples, characterized as (T350G) European variants. However, no signal of intratypic recombination event was identified within the E6–E7 region. Molecular and evolutionary analyses of HPV16 genomes from distinct geographical locations might provide valuable information about viral evolution and oncogenecity.

The prevalence of infection caused by different categories of diarrhoeagenic E. coli (DEC) strains, including enteroaggregative (EAEC), enteropathogenic (EPEC), enterotoxigenic (ETEC), enteroinvasive (EIEC) and enterohaemorrhagic (EHEC) E. coli, in children who suffered from diarrhoea (n = 3943) or did not have diarrhoea (n = 1026) were analysed in two areas in Bolivia over a period of 4 years. We also analysed the seasonality of DEC infections and severity of diarrhoea in children with DEC infection and compared antibiotic resistance in DEC strains isolated from children with and without diarrhoea. Stool samples were analysed for the presence of DEC by culturing followed by PCR. The most prevalent DEC categories in samples from the children were: EAEC (11.2 %); ETEC (6.6 %); EPEC (5.8 %); and EIEC and EHEC (<1 %). DEC strains were isolated significantly more often from diarrhoea cases (21.6 %) than from controls (17.6 %; P = 0.002). The number of children with diarrhoea associated with EAEC, EPEC and ETEC infections peaked in the Bolivian winter (April–September), although the proportion of DEC-positive stool samples was higher during the warm rainy season (October–March). High levels of antibiotic resistance were detected among the DEC strains. In particular, resistance to tetracycline and sulfamethoxazole–trimethoprim was significantly higher in strains isolated from individuals with diarrhoea than in samples from controls. The severity of disease in children infected with EAEC, EPEC and ETEC varied from mild to severe diarrhoea, although disease severity did not differ significantly between the different DEC categories. ETEC, EPEC and EAEC are commonly found in Bolivia and may cause severe disease in children.

A total of 1878 non-duplicate clinical Escherichia coli isolates (comprising 1711 urinary isolates and 167 blood-culture isolates), which were collected from multiple centres in Hong Kong during 1996–2008, were used to investigate the prevalence and molecular epidemiology of plasmid-mediated fosfomycin (fos) resistance genes. Eighteen of the 1878 clinical E. coli isolates were fosfomycin resistant, of which six were fosA3 positive and two were positive for another fosA variant (designated fosKP96). No isolates had the fosC2 gene. The clones of the eight isolates were diverse: sequence type (ST) 95 (n = 2), ST118 (n = 1), ST131 (n = 1), ST617 (n = 1), ST648 (n = 1), ST1488 (n = 1) and ST2847 (n = 1). In the isolates, fosA3 and bla CTX-M genes were co-harboured on conjugative plasmids with F2:A−:B− (n = 2), N (n = 1), F–:A−:B1 and N (n = 1) and untypable (n = 2) replicons. Both fosKP96-carrying plasmids belonged to replicon N. RFLP analysis showed that the two F2:A−:B− plasmids carrying fosA3 and bla CTX-M-3 genes shared the same pattern. Complete sequencing of one of the two F2:A−:B− plasmids, pFOS-HK151325 (69 768 bp) demonstrated it to be >99 % identical to the previously sequenced plasmid pHK23a originating from a pig E. coli isolate in the same region. This study demonstrated the dissemination of fosA3 genes in diverse E. coli clones on multiple bla CTX-M-carrying plasmid types, of which F2:A−:B− plasmids closely related to pHK23a were shared by isolates from human and animal sources.

Carbapenem-resistant Enterobacteriaceae have been frequently reported worldwide. They represent a serious concern because of the limited therapeutic options. The aim of this study was to investigate the molecular epidemiology of 14 Klebsiella pneumoniae carbapenemase (KPC) producers among 345 clinical isolates of Enterobacteriaceae with reduced susceptibility to carbapenems recovered from 11 separate hospitals in southern Brazil. The bla KPC-2 gene was detected in 14 isolates (4 %): six Enterobacter cloacae, five K. pneumoniae and three Serratia marcescens. Most of these isolates exhibited high-level resistance against β-lactams and ciprofloxacin, while the most active drugs were polymyxin B and amikacin. Genetic environment analysis, based on the classical Tn4401 structure, revealed six distinct platforms. Plasmids carrying bla KPC-2 were not typable and most were approximately 20 kb. Only KPC carbapenemases were found among the isolates studied, highlighting the local relevance of these enzymes in acquired resistance to carbapenems in Enterobacteriaceae. Our results contribute to the understanding of carbapenem resistance in Enterobacteriaceae and to the molecular characterization of KPC-2-producing isolates in Brazil.

A total of 69 strains of Escherichia coli from patients in the Taizhou Municipal Hospital, China, were isolated, and 11 strains were identified that were resistant to bacitracin, chloramphenicol, tetracycline and erythromycin. These strains were PCR positive for at least two out of three genes, ybjG, dacC and mdfA, by gene mapping with conventional PCR detection. Conjugation experiments demonstrated that these genes existed in plasmids that conferred resistance. Novel ybjG and dacC variants were isolated from E. coli strains EC2163 and EC2347, which were obtained from the sputum of intensive care unit patients. Genetic mapping showed that the genes were located on 8200 kb plasmid regions flanked by EcoRI restriction sites. Three distinct genetic structures were identified among the 11 PCR-positive strains of E. coli, and two contained the novel ybjG and dacC variants. The putative amino acid differences in the ybjG and dacC gene variants were characterized. These results provide evidence for novel variants of ybjG and dacC, and suggest that multiple drug resistance in hospital strains of E. coli depends on the synergistic function of ybjG, dacC and mdfA within three distinct genetic structures in conjugative plasmids.

Streptococcus pneumoniae diseases are a rare but increasingly recognized trigger of atypical haemolytic uraemic syndrome (HUS) in young children and associated with a higher mortality rate than diarrhoea-associated HUS. This study aimed to determine the importance of neuraminidase A (NanA) and genomic diversity in the pathogenesis of pneumococcal HUS (pHUS). We investigated the nanA gene sequence, gene expression, neuraminidase activity and comparative genomic hybridization of invasive pneumococcal disease (IPD) isolates from patients with pHUS and control strains matched by serotype and sequence type (ST), isolated from patients with IPD but not pHUS. The nanA sequence of 33 isolates was determined and mutations at 142 aa positions were identified. High levels of diversity were observed within the NanA protein, with mosaic blocks, insertions and repeat regions present. When comparing nanA allelic diversity with ST and disease profile in the isolates tested, nanA alleles clustered mostly by ST. No particular nanA allele was associated with pHUS. There was no significant difference in overall neuraminidase activity between pHUS isolates and controls when induced/uninduced with N-acetylneuraminic acid. Comparative genomic hybridization showed little difference in genetic content between the pHUS isolates and the controls. Results of gene expression studies identified 12 genes differentially regulated in all pHUS isolates compared with the control. Although neuraminidase enzyme activity may be important in pHUS progression and contribute to pathogenesis, the lack of a distinction between pHUS isolates and controls suggests that host factors, such as acquired abnormalities of the alternative complement cascade in young children, may play a more significant role in the outcome of pHUS.

Anaerobic bacterial infection is often accompanied by abscess formation; however, few in vivo studies have been published with descriptive data specifically evaluating antimicrobial activity in the presence of abscesses. The aim of this study was to establish a murine model of anaerobic infection with abscess formation and to verify the utility of this model for evaluating the in vivo efficacy of an antimicrobial agent. A clinical isolate of Fusobacterium necrophorum was inoculated into the caudal vein of immunocompetent BALB/c mice at 108 c.f.u. per mouse. Changes in body weight, bacterial load and histopathology of key organs were evaluated. After inoculation, bacterial counts in the liver increased from 104 to 108 c.f.u. after 1–3 days, and liver abscess formation was observed on the day following infection. Abscess formation and bacterial growth were not observed in other organs. In this model, 3 days of treatment with 5 mg metronidazole kg−1 eradicated F. necrophorum in the liver; however, a reduction in bacterial load was not observed with 0.05 mg metronidazole kg−1. In this study, we established a novel murine model of F. necrophorum liver abscess via haematogenous infection that may be useful for investigating in vivo antimicrobial activity against anaerobic abscesses and understanding the pathogenesis of F. necrophorum infection.

In recent decades the incidence of Candida endocarditis has increased dramatically. Despite the application of surgery and antifungal therapy, Candida endocarditis remains a life-threatening infection with significant morbidity and mortality. We report a 37-year-old male drug abuser presenting with high fever, chest pain, loss of appetite and cardiac failure. His echocardiography revealed mobile large tricuspid valve vegetations. Fungal endocarditis was confirmed by culturing of the resected vegetation showing mixed growth of Candida albicans and Candida tropicalis, although three consecutive blood cultures were negative for Candida species. Phenotypic identification was reconfirmed by sequencing of the internal transcribed spacer (ITS rDNA) region. The patient was initially treated with intravenous fluconazole (6 mg kg−1 per day), followed by 2 weeks of intravenous amphotericin B deoxycholate (1 mg kg−1 per day). Although MICs were low for both drugs, the patient’s antifungal therapy combined with valve replacement failed, and he died due to respiratory failure.