Proteinase, lipase and α-glucosidase activity in 81 clinically isolated Candida strains grown in nutrient-restricted media were comparatively measured, and the correlation between inducible enzyme activity and fungal growth was analyzed. Enzyme activity was assayed by colorimetric methods. Extracellular proteinase activity was significantly higher in Candida albicans, followed by C. parapsilosis and C. tropicalis. Extracellular lipase activity was observed in all 6 Candida spp. in the descending order of C. albicans, C. tropicalis and C. parapsilosis. Cell surface α-glucosidase activity was significantly higher in C. tropicalis, C. albicans, and C. parapsilosis, but was not detected in the other three species. A relatively strong correlation was observed between proteinase activity and fungal growth (correlation coefficient: 0.72); a mild correlation was observed between lipase activity and fungal growth (0.55); and a slight correlation was obserbed between α-glucosidase activity and fungal growth (0.32). There was no correlation among the strains in the activity levels of the three enzymes. These results suggest that pathogenic fungi produce larger amounts of inducible hydrolytic enzymes, and that proteinase is most likely to be related to fungal growth in nutrient-restricted conditions.

The membrane-spanning domain (MSD) of HIV-1 envelope protein (Env) has an additional glycine residue within a well-conserved putative transmembrane helix-helix interaction motif, GXXXG, and forms a G⁶⁹⁰ G⁶⁹¹ XXG⁶⁹⁴ sequence (G, glycine; X, any residues; the numbering indicates the position within the Env of an infectious molecular clone, HXB2). Different from vesicular stomatitis virus G (VSV-G), the glycine residues of the GXXXG motif of HIV-1 showed higher tolerance against mutations, and a simultaneous substitution of G690 and G694 with leucine residues only modestly decreased fusion activity and replication capacity of HIV-1. When G691 was further substituted with alanine, phenylalanine or leucine residue while G690 and G694 were substituted with leucine residues, the efficiency of membrane fusion decreased, with the decrease greatest occurring with the leucine substitution, a less severe decrease with phenylalanine, and the least severe decrease with alanine. Substitution with leucine residue also decreased the incorporation of Env onto virions, and the mutant showed the most delayed replication profile. Thus the presence of the extra glycine residue, G691, may increase the tolerance of the other two glycine residues against mutations than VSV-G. The fact that a more severe defect was observed for the leucine residue than the phenylalanine residue suggested that the function of Env depended on the steric nature rather than on the simple volume of the side chain of the amino acid residue at position 691. Based on this result, we propose a hypothetical model of the association among MSDs of gp41, in which G⁶⁹¹ locates itself near the helix-helix interface.

During the febrile illness epidemic in Bangladesh in 2002, 58 people died out of the 6,132 affected. Two hundred hospitalized patients were analyzed clinically, serologically and virologically to determine the features of this dengue infection. Among the 10- to 70-year-old age group of the 200 clinically suspected dengue patients, 100 (50%) were confirmed as dengue cases by virus isolation and dengue IgM-capture ELISA. Of the 100 dengue-confirmed cases, the mean age was 29.0 (±12.4). The possible dengue secondary infection rate determined by Flavivirus IgG-indirect ELISA was 78% in 2002. Eight dengue virus strains were isolated, representing the first dengue virus isolation in the country, and all of the strains were dengue virus type-3 (DEN-3). Sequence data for the envelope gene of the DEN-3 Bangladeshi isolates were used in a phylogenetic comparison with DEN-3 from other countries. A phylogenetic analysis revealed that all 8 strains of DEN-3 were clustered within a well-supported independent sub-cluster of genotype II and were closely related to the Thai isolates from the 1990s. Therefore, it is likely that the currently circulating DEN-3 viruses entered Bangladesh from neighboring countries.

We analyzed protein kinase R (PKR)-binding domain sequences of hepatitis C virus (HCV) NS5A protein and the profile of HCV-specific antibodies from pretreatment sera of HCV-chronically infected patients. Results were compared with clinical data to verify their influence on the course and result of therapy. Of 9 patients enrolled in a 12-month treatment with pegylated interferon alpha (PEG-IFN-α) plus ribavirin (RBV), 6 patients responded to therapy, as assessed by the lack of HCV RNA in their sera, and 3 did not. Among 8 HCV-1b-infected patients, those who responded did not have significantly more mutations in the IFN sensitivity determining region (ISDR) compared to non-responders (P = 0.637). Similarly, in the remaining 26-amino acid region of the PKR-binding domain, behind ISDR, the number of mutations did not differ significantly between the two groups (P = 0.796). A correlation was found between the presence of envelope 2 (E2)-specific antibodies and the result of treatment (P = 0.048). This pilot study indicates that mutations in the PKR-binding domain of HCV genotype 1b do not correlate with outcome of PEG-IFN-α/RBV therapy. However, the presence of E2-specific antibodies in the pretreatment sera of HCV-chronically infected individuals could serve as a prognostic marker predicting the result of treatment, before its initiation.

Three 80- to 95-month-old Holstein dairy cattle infected naturally with the agent of bovine spongiform encephalopathy (BSE) and slaughtered at abattoirs in Japan were examined for the distribution of disease-specific and protease-resistant prion protein (PrP^Sc) by immunohistochemistry (IHC) and Western blot (WB) analyses. The cattle showed no clinical signs or symptoms relevant to BSE but were screened as positive by enzyme-linked immunosorbent assay, a rapid test for BSE. This positive result was confirmed by IHC or WB in a specimen of the medulla oblongata. Histopathologically, these cattle showed no vacuolation in tissue sections from the central nervous system except for the medulla oblongata. Both IHC and WB analyses revealed PrP^Sc accumulation in the brain, spinal cord, satellite and ganglionic cells of the dorsal root ganglia, and the myenteric plexus of the distal ileum. In addition, small amounts of PrP^Sc were detected in the peripheral nerves of 2 cattle by WB. No PrP^Sc was demonstrated by either method in the Peyer’s patches of the distal ileum; lymphoid tissues including the palatine tonsils, lymph nodes, and spleen; or other tissues. The distribution of PrP^Sc accumulation in the preclinical stage was different between naturally infected cattle and cattle inoculated experimentally with the BSE agent.

Vibrio vulnificus is a Gram-negative bacterium which is associated with severe infections in humans. We experienced two cases of sucrose-fermenting V. vulnificus infection. The causative agents in both cases were unidentifiable by conventional identification systems because of their unique characteristics, and sequencing of 16S rDNA was found to be useful for diagnosis.

To investigate Haemophilus influenzae type b (Hib) infection in Vietnamese children under the age of 5 years, cerebrospinal fluid (CSF) samples from patients with meningitis were screened for Hib, and isolates were subjected to evaluation of susceptibility to 12 antibiotics, biotyping, and genotyping with pulsed-field gel electrophoresis (PFGE). The major biotype was type II (68.3%), followed by type I (22.8%). Among 79 Hib isolates, 45 (57%) were β-lactamase-producing and ampicillin-resistant (44 and 1 isolates produced TEM-1- and ROB-1-type β-lactamases, respectively), and 34 isolates (43%) were β-lactamase-nonproducing and ampicillin-sensitive. No β-lactamase-nonproducing and ampicillin-resistant isolates were found. The PFGE patterns of Hib isolates were highly divergent, but most could be classified into three clusters. We also investigated Hib colonization in household contacts of patients, and found that Hib isolates from the CSF of patients and from nasopharyngeal cavities of household contacts showed the same PFGE patterns. This observation suggested that household contacts of patients are a possible reservoir of Hib.

The aim of this study was to evaluate the value of nalidixic acid resistance as an indicator of decreased susceptibility to ciprofloxacin (MIC = 0.125 - 1 mg/L) in Salmonella isolates from humans (n = 620) in Turkey. One isolate was found to be resistant, and the remaining isolates were susceptible to ciprofloxacin with the Clinical and Laboratory Standards Institute breakpoints; however, 75 isolates (12.1%) had decreased susceptibility. Resistance to nalidixic acid was observed in 76 (12.3%) isolates in the disk diffusion test. Seventy-four of these isolates had decreased susceptibility, one was fully resistant, and one isolate was susceptible to ciprofloxacin. One isolate with decreased susceptibility to ciprofloxacin was intermediate to nalidixic acid. Screening with 30- μg nalidixic acid disks had a sensitivity of 98.6% and a specificity of 99.8% for determination of decreased susceptibility to ciprofloxacin.

Pantoea infections are uncommon in humans. Most reports have involved adults or children after thorn injuries. There are only a few reports of systemic infections with Pantoea. This is the first report of the clinical picture of systemic Pantoea spp. infection in neonates as observed during an outbreak in a neonatal intensive care unit caused by infected parenteral nutrition solutions. Even though detected early, the infections had a fulminant course, causing septicemic shock and respiratory failure. Pulmonary disease was prominent and presented mainly as pulmonary hemorrhage and adult respiratory distress syndrome. The organism was sensitive to most antibiotics used in neonatal intensive care units, but the clinical response to antibiotic therapy was poor. The fatality rate was very high: 7 out of 8 infected infants succumbed to the infection (87.5%).

Drug-resistance genotypes were investigated in a patient under treatment with anti-HIV drugs. Since the drug resistance-associated mutations in plasma HIV-1 RNA and proviral DNA in peripheral blood mononuclear cells (PBMCs) were inconsistent, changes were followed over time, and the discrepancy was shown to persist for a long period. In plasma HIV-1 RNA, D67N, K70R, T215Y, and Y188L were present in the reverse transcriptase (RT) region, and two primary mutations, I84V and L90M, were noted in the protease (Pro) region. In contrast, in proviral DNA, no drug resistance-associated mutations were found in the RT region, and mutations such as L90L/M were only infrequently present in the Pro region. This situation persisted for more than 3 years. In addition, sequencing analysis of the V3 loop in the envelope gene showed that non-syncytium-inducing/macrophage-tropic viruses contribute to acquisition of drug resistance. In this study, drug-resistant viruses were produced primarily at macrophages, and drug-sensitive viruses were maintained in PBMCs as a reservoir.

A case of deep brain abscess by Gemella morbillorum is described. Due to high fever, lethargy, severe headache, and the risk of intraventricular rupture of the suppurative lesion, a CT-guided stereotactic aspiration of the abscess was successfully performed. The patient responded well to a 6-week course of meropenem, metronidazole, and fluconazole. Gemella spp. should not be considered as trivial commensals of the mucous membranes, but appear as emerging pathogens involved in endocarditis, septic shock, and necrotizing pneumonia, as well as in serious intracranial infections.

Trichosporon fungemia is usually seen in neutropenic patients with underlying hematological malignancies. In this report we describe a fatal case of Trichosporon asahii fungemia in a non-neutropenic patient with a non-hematological malignancy. For 1 week the patient exhibited hematuria, weakness, easy fatigability and headaches. At admission she had anemia, renal failure and evidence of right hydronephrosis and bladder wall masses as detected by CT scan. She did not have a history of tobacco abuse, contact with urinary carcinogens or Schistosoma infestation; her clinical picture was suggestive of bladder cancer. After some investigations the patient underwent radical cystectomy and ileal conduit surgery because of transitional cell carcinoma in the urinary bladder. After an initial uneventful improvement postoperatively the patient deteriorated and died of septic shock despite all reanimation efforts and antibiotherapy including fluconazole. The blood culture obtained 4 days before the patient died revealed T. asahii, which was isolated on the day she died and found to be resistant to fluconazole and caspofungin. This report suggests that clinicians remain aware that T. asahii fungemia may develop in clinically deteriorated patients even if they do not have a hematological malignancy.

Our report highlights a case of severe childhood salmonellosis related to a pet turtle, a red-eared slider (Trachemys scripta elegans). A 6-year-old girl had gastroenteritis complicated with sepsis caused by serotype Paratyphi B, which shared the same pulsed-field gel electrophoresis profiles with the organism isolated from a pet turtle. Based on our literature survey on childhood invasive salmonellosis acquired from reptiles, this case is the first documented reptile-associated salmonellosis including sepsis caused by this serotype.