In this study we aimed to learn about the nature and frequency of katG, inhA, and rpoB gene mutations underlying isoniazid (INH) and rifampin (RMP) resistance in clinical Mycobacterium tuberculosis complex isolates. The Silver Sequence DNA sequencing method was used to detect the resistance condition of 22 INH, 6 RMP, and 13 INH and RMP in previously determined drug-resistant clinical M. tuberculosis isolates. Thirty of 35 (85.7%) INH-resistant strains and 14 of 19 (73%) RMP-resistant strains were found to have a mutation in the analyzed katG gene fragment or inhA locus and rpoB gene fragment. In the katG gene region, the codons of mutation detected were determined to be 315 (23 of 30, 76.6%), 279 (4 of 30, 13.3%) and 293 (1 of 30, 3.3%), a finding that has not been reported previously. Our findings demonstrated that the most frequent mutation pattern was Ser315Thr at codon 315 with a rate of 60% (18 of 30). In 5 (16.6%) isolates, a nucleotide change was detected which is associated with INH resistance from –15^th C to T in the inhA locus. In the rpoB gene region, codons possesing point mutations were 531 (9 of 14, 64.2%), 516 (1 of 14, 7.1%), 524 (1 of 14, 7.1%), and 545 (4 of 14, 28.6%), which has not been reported previously. We believe about that our present study supplies important data on the different kinds of mutations occurring at various target loci for associated RMP and INH resistance in clinical isolates of our restricted region.

With the increasing resistance of group A streptococci (GAS) to macrolides in some countries including Korea, throat cultures were taken from healthy elementary school children to isolate GAS in 2004. Antibiotic susceptibility, macrolide-resistant phenotypes and genotypes as well as emm genotyping were studied and compared with previous data in 2002. In the present study, resistance rates to erythromycin and clindamycin in 2004 decreased to 9.8 and 8.8%, respectively, compared with 51.0 and 33.7%, respectively, in 2002. While emm44/61 increased from 0 to 29.3%, emm12 decreased from 34.4 to 6.4% during the 2002 - 2004 period. All emm44/61 strains were susceptible to erythromycin, while 81.0% of emm12 strains were resistant to erythromycin. The dramatic decrease of erythromycin resistance during this short period might be related to a change in the distribution of emm types in the community.

Tuberculosis is still a severe public health issue in eastern Asia, and Sichuan is the key area for tuberculosis control in China. To determine the phenotypic and mutation patterns of drug resistance in Mycobacterium tuberculosis isolates from Sichuan, the drug susceptibility of 198 clinical isolates was examined. Among these isolates, 76 drug-resistant and 20 susceptible isolates were analyzed for the rpoB, embB, and katG and inhA regulatory regions. These are mutations believed to associate with rifampin (RIF), ethambutol (EMB), and isoniazid (INH) resistance, respectively. Of the 60 RIF-resistant isolates, 54 (90.0%) carried mutations on the amplified fragment of the rpoB gene, and the most common one (64.8%, 35/54) was at codon 531. Two new mutation patterns were recognized: one isolate harbored three mutations at codons 511, 516, and 518, and the other carried the dual mutation GAC→ACC at codon 516. A total of 30 INH-resistant isolates (60.0%, 30/50) had mutations at codon 315, whereas 4 (8.0%) had mutations at the inhA regulatory region. Among the 46 EMB-resistant isolates, 22 harbored the Met306 mutation. The results showed geographical variation in the mutation types of drug-resistant genes in M. tuberculosis isolates from Sichuan; this finding is valuable for the development of targeted and rapid molecular diagnostic methods suitable for specific regions.

During a study examining causes of diarrhea from May 2004 to May 2005, 808 stool specimens were collected from patients with acute diarrhea in Tehran. Fecal samples were cultured and identified according to the standard biochemical methods. Molecular identification of enteropathogens was carried out by amplification of their virulence genes by polymerase chain reaction. A total of 369 (45.6%) bacterial pathogens were recovered from 808 patients as follows: Shigella spp., 155 (45.6%); diarrheagenic Escherichia coli 143 (38.8%); Salmonella spp., 51 (13.8%); and Campylobacter spp., 20 (5.4%). Most of the diarrheagenic E. coli were Shiga toxin-producing E. coli, with 64 (44.7%) isolates, followed by 47 (32.9%) enterotoxigenic E. coli isolates; among Shigella spp. isolates, 69 (44.5%) Shigella flexneri were predominant. The molecular diagnosis of enteropathogens yielded a more accurate characterization of the prevalence of diarrhea-causing bacterial strains in Iran. The present study revealed a high prevalence of Shigella and diarrheagenic E. coli as the predominant causes of bacterial diarrhea in this region of the world. These two types of bacteria should therefore be considered when designing preventive strategies for people living in Iran.

Multiple drug-resistant strains of Acinetobacter have created therapeutic problems worldwide. This study was conducted to determine the antimicrobial susceptibility patterns and prevalence of bla_OXA-type carbapenemases among isolates of Acinetobacter spp. obtained from Iranian patients. Here, 128 Acinetobacter isolates were identified at the species level, and their susceptibilities to different antibiotics were determined using disk agar diffusion testing. Isolates were then subjected to multiplex-PCR targeting bla_OXA genes. More than 50% of the isolates showed multidrug resistance to different antibiotics. The rates of susceptibility to imipenem, meropenem, piperacillin-tazobactam, and amikacin were 50.7, 50, 42.1, and 38.2%, respectively. The MICs of carbapenems for the resistant isolates ranged from 64 to ≥256 μg/ml. All strains of Acinetobacter baumannii possessed a bla_OXA-51-like gene. The co-existence of bla_OXA-51-like/bla_OXA-23-like and bla_OXA-51-like/bla_OXA-24-like was detected in 25% (n = 32) and 17.9% (n = 23) of the isolates, respectively. Over 70% of carbapenem-resistant strains contained at least two genes encoding OXA-type carbapenemase. Resistance to carbapenems in the population of Acinetobacter strains in Iran is high, with the majority of isolates showing multidrug resistance. A wide diversity of OXA genes exists among the strains of A. baumannii in Iran. Detection of bla_OXA-51-like can be used as a simple and reliable method to differentiate A. baumannii strains from other species.

A procyanidin-rich extract from French maritime pine, Pycnogenol^® (PYC), is known as an antioxidant that exerts a variety of physiological activities and is widely used in human beings. We report here that PYC inhibits not only human immunodeficiency virus type-1 (HIV-1) binding to host cells, but also its replication after entry in susceptible cells in vitro. Prominent biochemical alterations induced by PYC were the elevated expression of an intracellular antioxidant protein, manganese superoxide dismutase (Mn-SOD), and the inhibition of phosphorylation of the ribosomal S6 protein. Interestingly, ectopic expression of Mn-SOD inhibited HIV-1 replication as well. Inhibition of HIV-1 replication associated with induced expression of Mn-SOD in cells treated with PYC suggests the potential of this natural antioxidant inducer as a new anti-HIV-1 agent.

Of 59 clinical isolates of Enterobacter cloacae from a teaching hospital in Sichuan, China, 18 isolates were shown to be resistant to oxyimino cephalosporins and aztreonam. Enterobacterial repetitive consensus PCR revealed that these isolates comprised 7 distinct genotypes. The presence of plasmids in the 18 clinical isolates was revealed by conjugational transfer of plasmids from E. cloacae to Escherichia coli with the further isolation of the plasmids in the transconjugants. Subsequent nucleotide sequencing and β-lactamase isoelectric focusing indicated that the plasmids encoded blaSHV, blaCTX-M and/or blaTEM genes, including genes for CTX-M-22 (13 strains), TEM-1 (12 strains), TEM-29 (1 strain), TEM-141 (1 strain), TEM-157 (1 strain), SHV-5 (1 strain), SHV-12 (1 strain), and SHV-70 (1 strain). The widespread presence of extended-spectrum β-lactamases in E. cloacae isolated from the southwest of China was likely due to the dissemination of resistance plasmids with the predominant genotype of blaCTX-M-22.

Mosaicism in vacA alleles with two distinct families of vacA signal sequences (s1 and s2) and two distinct families of middle region alleles (m1 and m2) has been reported. Research suggests that the vacA s1 genotype is closely associated with duodenal ulcer disease and with high cytotoxin production. The aims of this study were to evaluate the role of vacA genotyping with respect to gastric inflammation and injury, and clinical presentation in Iranian populations. Genomic DNA of biopsy specimens from patients with gastritis, peptic ulcer disease (PUD), or gastric cancer (GC) were characterized based on ureC (glmM), cagA, and vacA genotyping by using polymerase chain reaction. Of 167 patients including 33 with PUDs, 129 with non-ulcer dyspepsia (NUD), and 5 with GC, 96 (57.5%) cases were infected by Helicobacter pylori. Among these patients, H. pylori were isolated from 19 (57.7%) PUD patients, 74 (68.7%) NUD patients, and 3 (60%) GC patients. The cagA was detected in 76% of H. pylori-positive cases. The vacA s1-m2 genotype was the most prevalent in 7/19 PUD (37%) and 30/74 NUD (40.5%) patients with H. pylori infection. The prevalence of vacA s2-m1 (8%) was high in Iranian isolates. A significant association was not found between H. pylori genotypes and clinical outcomes. The vacA genotypes and cagA status were not useful markers for gastroduodenal diseases in Tehran, Iran.

Time-lapse video microscopic observation is useful for analysis of cell biology, especially in rapid response of immune cells. Dendritic cells (DCs) have multiple functions in the immune system, and DCs in peripheral blood play an especially important role at the front line of infection. We have developed a time-lapse video microscopic method for the evaluation of single myeloid DCs (MDC-1) from human peripheral blood. MDC-1 displayed generalized plasma membrane ruffling and phagocytosis of Pseudomonas aeruginosa. The morphological changes of MDC-1 increased following stimulation with P. aeruginosa but not after stimulation with supernatant from a P. aeruginosa culture. The activation of these morphological changes in MDC-1 could be quantitatively analyzed using the time-lapse video microscopy. This novel system may be useful for the evaluation of rapid response with human immune cells against bacterial infection.

Dengue hemorrhagic fever and dengue shock syndrome, the major life-threatening outcomes of severe dengue disease, are the consequence of plasma leakage in the vascular areas. We previously demonstrated that dengue virus (DV)-infected dendritic cells (DC) trigger vascular leakage through matrix metalloproteinase (MMP)-9 overproduction, however little is known concerning the consequences of direct infection of macrovascular endothelial cells (MVEC) by DV. In this study, we show that infection of primary human MVEC results in overproduction of MMP-2 and to a lesser extent of MMP-9, leading to enhanced endothelial permeability. This permeability was associated with loss of expression of the vascular endothelium-cadherin cell-cell adhesion. The MMP response to DV infection is strikingly different between DC and MVEC. Therefore, our results demonstrated that endothelial cells are an important target for DV infection, and that DV-induced MMP-2 overproduction by direct infection of endothelial cells may contribute to the pathogenesis of severe dengue infection.

To determine the prevalence and antimicrobial susceptibility of Streptococcus pneumoniae in the oropharynx of healthy children, throat swabs were obtained from 683 children and cultured. The disk diffusion method and the E test were used to test the antimicrobial susceptibility of the isolated organisms. Twenty-nine children (4.2%) harbored S. pneumoniae in their oropharynx. Fifteen (51.7%) of the isolates showed intermediate resistance to penicillin and 14 (48.3%) were susceptible. All strains were susceptible to rifampicin and moxifloxacin. One was resistant to telithromycin. The rates of resistance to clindamycin, erythromycin, chloramphenicol and tetracycline were 41.3, 44.8, 34.4, and 44.8%, respectively. Risk factors for S. pneumoniae carriage were also assessed.

It has not been well elucidated whether patients with measles who have undergone postexposure prophylaxis (PEP) exhibit milder clinical symptoms than would be expected in the absence of PEP. In the present study, we compared the severity of measles of patients who had undergone incidental PEP to those of patients who had not received such prophylaxis. An outbreak of measles occurred among the personnel of the Japan Self-Defense Forces (SDF) between 7 May and 14 June 2007 at 3 camps in or near Sendai City. All patients were admitted to SDF Sendai Hospital for isolation and care. Measles was confirmed in 21 patients (age: 21.9 ± 5.9 years). Of the 15 ill recruits (persons who had just joined SDF), 8 underwent incidental PEP on 10 May, 3 days after the onset of the prodromal symptoms in the first patient. The vaccination was not originally intended as PEP and was administered within the framework of a routine vaccination program. Compared to recruits without PEP (n = 7), the admission period was 5.6 days shorter, the period with high fever (39℃ or more) was 2.7 days shorter, and the maximum body temperature was 1.1℃ lower in the recruits with PEP (n = 8). These results suggest that PEP ameliorates the clinical symptoms of measles.

From May 2004 to October 2006, a prospective study was carried out in Dakar, Senegal, to update information about the antimicrobial susceptibility of Shigella spp. isolated from stool specimens. Among the 165 non-duplicate strains collected, 81 (49%) were identified as Shigella flexneri, 75 (45%) as Shigella sonnei, 5 (3%) as Shigella boydii, and 4 (2%) as Shigella dysenteriae. Disk diffusion testing revealed that the majority of isolates were resistant to sulphonamides, trimethoprim-sulfamethoxazole, streptomycin, and tetracycline (respective overall resistance rates: 90, 90, 96, and 94%). More than half of the S. flexneri isolates were resistant to amoxicillin, amoxicillin-clavulanic acid, and chloramphenicol (respective resistance rates: 59, 58, and 52%), and almost all of the S. sonnei isolates were susceptible to these antimicrobials (respective resistance rates: 4, 1, and 4%). Only one isolate (belonging to the species S. sonnei) was resistant to nalidixic acid and displayed reduced susceptibility to ciprofloxacin.

The aim of this study was to search for three plasmid-encoded, quinolone-resistant determinants: qnrA, qnrB, and qnrS. Thus, 460 Gram-negative strains belonging to 11 different genera (clinical, 347; non-clinical, i.e., from a rectal swab, 113), of which 40% were ciprofloxacin-resistant, were recovered from patients in an intensive care unit at the Istanbul Medical Faculty, Turkey, in the years 2000 and 2006. PCR with primers specific for qnrA, qnrB, and qnrS genes and primers specific for a series of ESBL genes were used. One qnrB1 and two qnrS1 genes were identified in three ESBL-positive isolates, whereas no qnrA-positive strain was found. The qnrB1 determinant was identified in a ciprofloxacin-susceptible Enterobacter cloacae isolate that expressed CTX-M-15 β-lactamase. Two qnrS1-determinants were found in two ciprofloxacin-susceptible E. cloacae isolates that were clonally related, but that had been isolated from different patients; both of these isolates expressed the same ESBL, CTX-M-3. This study detected the first plasmid-mediated quinolone-resistance determinants qnrB1 and qnrS1, among clinical strains obtained from patients in Turkey.

Salmonella enterica serovar Paratyphi A is a causative agent of paratyphoid fever. The clinical syndrome caused by paratyphoid fever overlaps with other febrile illnesses and cannot be distinguished from typhoid fever. Conventional methods used for diagnosis are time consuming, costly, and labor-intensive. We evaluated the specificity, sensitivity, and application of a multiplex polymerase chain reaction (PCR) previously developed by the method (Ou, H.Y., Teh, C.S.J., Thong, K.L., et al., J. Mol. Diagn., 9, 624-630, 2007) using 6 S. Paratyphi A, 22 S. Typhi, and 85 other Salmonella serovars as well as 36 non-Salmonella strains. The detection limit of the multiplex PCR was 4 × 10⁴ cfu ml^-1. In a blind test of the other 50 strains, this multiplex PCR correctly identified the only S. Paratyphi A in the panel of strains. The sensitivity of this PCR using spiked blood and stool samples was 1 × 10⁵ cfu ml^-1 and 2 × 10⁵ cfu ml^-1, respectively, but increased to 1 × 10⁴ cfu ml^-1 and 2 × 10³ cfu ml^-1 after 5-h enrichment. We believe that this multiplex PCR is a promising technique for the specific and sensitive detection of S. Paratyphi A in clinical, environmental, and food samples.

Rickettsial infection of cattle and sika deer from a pastureland in Hidaka District, Hokkaido, Japan was examined by serological and molecular methods. Serum samples from 8 of 83 (9.6%) cattle reacted with Rickettsia helvetica antigens in an IFA test, with titers ranging from 1:160 to 1:640, while serum samples from 15 of 22 (68.2%) deer were positive for R. helvetica, with titers ranging from 1:80 to 1:640. In a genus-specific nested PCR based on gltA, no cattle were positive for Rickettsia, while 14 of 22 (63.3%) samples obtained from deer tested positive. Sequence analysis revealed that positive samples from sika deer showed 100% nucleotide sequence identity with the known sequence of Rickettsia asiatica.

Post-sternotomy infections are a kind of nosocomial infection involving the mediastinum space and the sternum, with a high morbidity and mortality rate. The present study was carried out to identify the incidence of mediastinitis following cardiac surgery and the most common risk factors. Cardic patients undergoing surgery were studied for suspicious mediastinitis infection at the Madani Heart Center, Tabriz, Iran from 2004 to 2006. The most common isolated agents included the coagulase-negative staphylococci, Staphylococcus aureus, Pseudomonas aeruginosa and Enterobacter spp. Incidence of postoperative mediastinitis after cardiac surgery was 1.2%. The mortality rate of postoperative mediastinitis was high (34.3%). Wound infection, especially mediastinitis following cardiac surgery, is rare but could be life-threatening. The most important step in the management of wound infections is prevention, and preventive measures could be strengthened by identifying the risk factors.

Determining the local circulating strain of influenza is essential to prevent and control epidemics. In the years 2004 and 2005, the National Influenza Center of Thailand received 3,854 and 3,834 specimens, respectively, from patients throughout the country, including submissions from 4 established influenza surveillance sentinel sites. In 2004, of 539 influenza-positive specimens, 461 were positive for influenza A and 78 were positive for influenza B by isolation. Influenza A subtyping revealed that 249, 197, and 15 isolates were H1N1, H3N2, and H5N1, respectively. In 2005, of 748 influenza-positive specimens, 492 were influenza A and the remaining 256 were influenza B. The results of influenza A subtyping indicated that 55, 437, and 5 isolates were H1N1, H3N2, and H5N1. All isolated strains of subtype H1N1 were A/New Caledonia/20/99-like. The isolated strains of H3N2 were A/Fujian/411/2002-like in the first half of the year 2004, while those in the latter half of 2004 gradually drifted to a mixture of A/Wellington/1/2004-like, A/California/7/2004-like, and A/Wisconsin/67/2005-like, and this mixture continued through the end of 2005. The influenza B strains were B/Sichuan/379/99-like, B/Hong Kong/330/2001-like, B/Shanghai/361/2002-like and B/Malaysia/2506/2004-like. The strains circulating in the years 2004 and 2005 were antigenically similar to the vaccine formulas recommended in the same period by WHO. Our results underscore that local influenza surveillance plays an important role in responding to epidemics and potential pandemics.

In Nepal, hepatitis B and hepatitis C are considerable health problems. This study aimed to assess the trends of hepatitis B virus (HBV) and hepatitis C virus (HCV) seroprevalence in blood donors over the last 6 years nationwide and in the urbanized setting of Kathmandu Valley. This was a retrospective study conducted among Nepalese blood donors through the years 2001/2002-2006/2007. Serum samples were tested for hepatitis B surface antigen and anti-HCV antibodies using third generation ELISA tests. The donors' information was collected via the donor record register through their respective Blood Transfusion Services. The software, Winpepi ver 3.8 was used for statistical analysis. The overall seroprevalence rates of HBV and HCV in a nationwide analysis were observed to be 0.82 and 0.47%, respectively, and at Central Blood Transfusion Service (CBTS), Kathmandu, the rates were 0.92 and 0.71%, respectively. The seroprevalence of HBV was significantly higher than the seroprevalence of HCV, both nationwide and at CBTS (P < 0.05). An overall significantly decreasing trend was observed in HBV and HCV seroprevalence both nationwide and at CBTS, Kathmandu, over the last 6 years (P < 0.05). Though the overall trend was significantly decreasing, the test for departure from a linear trend also showed a statistically significant result (P < 0.05).