First characterization of Tn1546-like structures of vancomycin-resistant Enterococcus faecium Thai isolates.

INTRODUCTION: Vancomycin-resistant Enterococcus faecium (VREfm) carrying vanA was first isolated from patient at Siriraj Hospital, Thailand in 2004. Since then, VREfm isolates have been detected increasingly in this 2500-bed university hospital. To understand the epidemiology of vanA VREfm in this setting, the isolates collected during 2004-2013 were characterized. METHODS: A total of 49 vanA VREfm isolates previously confirmed by multiplex PCR were characterized by determining resistance phenotypes to vancomycin, teicoplanin, ampicillin and ciprofloxacin by broth microdilution method. Multilocus sequence typing (MLST) and virulence genes of those isolates were investigated. The Tn1546 structure diversity was studied by long-range overlapping PCR and primer walking sequencing. RESULTS: Of all isolates studied, 9 sequence types (ST17, ST80, ST78, ST730, ST203, ST18, ST280, ST64, ST323) in clonal complex 17 and a novel ST1051 were revealed. The esp-positive isolates were 73.5%. Of all vanA operons characterized, at least 9 types of Tn1546-like structures were detected. All of vanA determinants contained 5'-end different from the Tn1546 prototype. Approximately 47% of them also carried the insertion sequence IS1251 at the intergenic region between vanS and vanH. Interestingly, another IS (ISEfa4) was found to be inside the sequence of IS1251 in ST17 isolate. CONCLUSION: Heterogeneity of vanA VREfm was observed. Nearly all of isolates studied belonged to CC17. One novel ST1051 strain was detected. Isolates in the initial period carried vanA operon similar to the prototype. The diversity of vanA determinants has been increased in the recent isolates. A novel vanA operon structure was detected.

The usefulness of Neisseria gonorrhoeae strain typing by Pulse-Field Gel Electrophoresis (PFGE) and DNA detection as the forensic evidence in child sexual abuse cases: a case series.

Diagnosis of alleged child sexual abuse can be made from history in conjunction with physical examination, psychosocial evaluation, and laboratory investigations. Sexually transmitted infection associated with sexual abuse is found in 5 % of the victims, with Neisseria gonorrhoeae being the most common organism. Identification of sexually transmitted disease, particularly N. gonorrhoeae infection, can be useful for the diagnosis of sexual abuse and thus, the initiation of the child protection process. Polymerase Chain Reaction (PCR) is a newer diagnostic assay with a higher sensitivity compared with conventional culture method. In addition, N. gonorrhoeae strain typing can also be used to identify the abuser. In this case series, we present the application of N. gonorrhoeae strain typing (PFGE technique) to identify the abuser, and the confirmation of gonococcal vaginitis by PCR technique.

First case report of brucellosis in a child in Thailand.

Brucellosis is uncommon in children. In Thailand, there have been only seven adult cases reported, all with Brucella melitensis. We describe here the first reported pediatric case of brucellosis in Thailand. A 12-year old boy presented with prolonged fever for one month, pancytopenia, pneumonia and peritonitis. The blood culture grew out Brucella melitensis. He responded well to combination therapy consisting of doxycycline and gentamicin. He recovered fully without relapse during the 6 month follow-up.

Clinical characteristics and outcomes of acute bacterial meningitis in adults at a tertiary university hospital in Thailand.

The epidemiology of acute bacterial meningitis varies among settings, areas, and times. This study aimed to determine the clinical characteristics, the causative organisms and their antibiotic susceptibility, and the outcomes of patients with acute bacterial meningitis in Thai adults. This retrospective study included hospitalized patients aged ≥18 years diagnosed with acute bacterial meningitis at Siriraj Hospital during January 2002-December 2016. Of 390 patients, the median age was 48 years, 51% were male, and 80% had at least 1 underlying illness. Over half (54%) of patients had community-acquired bacterial meningitis (CBM), and 46% had nosocomial bacterial meningitis (NBM). The triad of acute bacterial meningitis (fever, headache, and neck stiffness) was found in 46% of CBM and 15% of NBM (P < .001). The causative organisms were identified in about half (53%) of patients. Most study patients (73%) received antibiotic treatment before the CSF collection. Causative organisms were identified more frequently in CBM (P < .001). Gram-positive cocci were more prevalent in CBM (P < .001), whereas Gram-negative bacilli were more common in NBM (P < .001). Streptococcus agalactiae (29%), Streptococcus pneumoniae (12%), and Streptococcus suis (11%) were most common in CBM, and all of them were sensitive to third-generation cephalosporins. Acinetobacter baumannii (19%), Klebsiella pneumoniae (16%), and Staphylococcus aureus (11%) were the most common organisms in NBM, and antibiotic-resistant isolates were frequently found among these organisms. Ceftriaxone monotherapy and meropenem plus vancomycin were the most common empiric antibiotic in CBM and NBM, respectively. Mortality was 19% among CBM and 23% among NBM (P = .338). HIV infection, alcoholism, pneumonia, shock, and disseminated intravascular coagulation were independent predictors of mortality. Mortality remains high among adults with acute bacterial meningitis in Thailand. A clinical practice guideline for acute bacterial meningitis should be developed that is based on local epidemiology and microbiology data.

Viability of Lactobacillus casei strain Shirota (LcS) from feces of Thai healthy subjects regularly taking milk product containing LcS.

OBJECTIVE: To demonstrate the recovery of Lactobacillus casei strain Shirota (LcS) from feces of Thai subjects who regularly took LcS containing milk product for 1 week and demonstrate the disappearance of LcS after stopped taking milk product. MATERIAL AND METHOD: First fecal samples were collected from 20 healthy adults at 10 days after they abstained from all lactobacillus containing milk products. Second specimens taken after the subjects ingested LcS containing milk product for 7 days and third specimens at 7 days after they stopped taking LcS containing milk product. All the fecal specimens were culture for LcS using LLV-FOS culture medium and enumeration of LcS was calculated. All stool samples were also tested for the presence of LcS by using nested PCR to confirm the presence of LcS obtained from culture method. RESULTS: Both culture and nested PCR method showed that all the stools samples obtained from subjects prior to the administration of LcS containing milk product were devoid of LcS, except for 3 specimens which showed weakly positive test for PCR. At 7 days after ingesting LcS containing milk product, all stool specimens were positive for LcS on both culture and PCR method. At 7 days after stopped taking LcS containing milk product, 1/19 specimens were positive from culture and 6/ 19 specimens were positive for PCR method. CONCLUSION: LcS could survive in the gastrointestinal tract of Thai subjects and could be recovered from the feces after ingestion.

Chronic recalcitrant bacterial infection in steroid modified interstitial (stromal) keratitis: presentation and management.

OBJECTIVE: To report histopathologically proven bacterial infection manifested multifocal interstitial (stromal) keratitis (IK) with definite previous history of prolong topical steroid use. Standard managements of bacterial keratitis did not provoke enough benefit. MATERIAL AND METHOD: A retrospective analysis of 19 eyes in 15 patients referred to Siriraj Hospital between 2004 and 2010. RESULTS: Multifocal intrastromal infiltration, with relatively quiet ocular reaction and mild inflammation were initially presented in all eyes. They all previously had been diagnosed of presumed viral keratitis, and had been given topical corticosteroid treatment for a prolonged period of time without healing. Autoimmune disease workups were all negative. Corneal scrapings showed negative culture results in all eyes. However, bacteria within stromal lamellae with absent or minimal inflammatory cells were demonstrated in all eyes by corneal biopsies. In addition, cytology results obtained from 16S rDNA sequencing revealed Stenotrophomonas maltophilia in one eye and coagulase-negative staphylococci in two eyes. No case responded well to intensive topical and systemic antibiotics. However they were successfully treated with penetrating keratoplasty (11 eyes, 57.9%) or intrastromal antibiotic injections (8 eyes, 42.1%). CONCLUSION: Bacterial infection should be a concern in prolonged chronic IK. This was considered as primary bacterial IK or bacterial superinfection in immunocompromised cornea. Early recognition and appropriately aggressive managements contribute to successful outcome. Corneal biopsy is always essential and 16S rDNA sequencing is useful in this distinct clinical entity.

Incidence of invasive Group B Streptococcus (iGBS) infections and the factors associated with iGBS mortality in adults during 2013-2017: a retrospective study at Thailand's largest national tertiary referral center.

OBJECTIVES: To determine the incidence of invasive Group B streptococcal (iGBS) diseases and the factors significantly associated with iGBS mortality in adult patients. MATERIAL AND METHODS: This retrospective study included adults with a positive culture for GBS isolated from a sterile site at Siriraj Hospital - Thailand's largest tertiary care hospital - during January 2013 to December 2017. RESULTS: Of the 224 included patients, 75.9% had bacteraemia. The median age of patients was 63 years (interquartile range [IQR]: 53-73) and 52.7% were female. Among the 80% of all patients with comorbid diseases, diabetes mellitus (38.8%), cancer (18.8%), and heart disease (12.5%) were the most common. Skin and soft tissue infection (30.8%), septic arthritis (21.4%), primary bacteraemia (21.0%), and meningitis (7.1%) were the most common manifestations of iGBS diseases. The overall 30-day mortality was 11%. Patients that died were older and had more chronic kidney disease, bacteraemia, urinary tract infection, pneumonia, and iGBS-related morbidities than survivors. Pneumonia was the only factor independently associated with 30-day mortality with an adjusted odds ratio of 24.96 (95% confidence interval [CI]: 5.95-104.75). CONCLUSIONS: Invasive GBS is not uncommon in non-pregnant adults, particularly among older adults and those with diabetes. Concomitant bacteraemia was frequently observed in iGBS patients. The overall mortality was low, but significant morbidities were observed.KEY MESSAGESIn our study, iGBS was not uncommon among older adults and those with diabetes.Two-thirds of patients with iGBS had bacteraemia, and the overall 30-day mortality was 11%.

Association between adenoid bacteriology and clinical characteristics of adenoid-related diseases in children.

INTRODUCTION: The aim of this research is to find the association between the pathogenic bacteria obtained from the adenoid culture and clinical characteristics of adenoid-related diseases in children. METHODS: In this retrospective study, we reviewed the medical records of children who had adenoidectomy for adenoid-related diseases. Demographic data, diagnoses, indications for adenoidectomy and bacterial culture results were collected. The adenoid size was measured in the lateral skull X-ray as adenoid-nasopharyngeal ratio. Associations between the culture results and the demographic data, adenoid size, and the diagnoses were analyzed. RESULTS: There were 407 children who had adenoidectomy for obstructive sleep-disordered breathing (75.2%), otitis media with effusion (19.2%), and chronic sinusitis (5.6%). Median age was 5.9 years. Common pathogenic bacteria in the adenoid were Haemophilus influenzae (26.2%), Staphylococcus aureus (23.5%), Streptococcus pneumoniae (18.2%), and Moraxella catarrhalis (12%). The patient's age had significant association with the prevalence of pathogenic bacteria. S. pneumoniae was most prevalent in young children up to 7 years. S. aureus was more common in children over 7 years. H. influenzae had similar prevalence in all age groups. Size of the adenoid and type of adenoid-related diseases had no association with the outcome of bacterial culture. CONCLUSION: Age of the patients was the significant factor associated with the bacteriological findings of the adenoid while size and types of adenoid-related diseases were not associated with the outcome of bacterial culture.

Multiplex PCR for detection of clarithromycin resistance and simultaneous species identification of Mycobacterium avium complex.

Multiplex PCR (mPCR) was established for the simultaneous detection of clarithromycin (CLR) resistance and species identification of Mycobacterium avium complex (MAC). mPCR was tested on 218 MAC clinical isolates. CLR-resistance was detected by mPCR in 31 of 35 isolates identified by a microdilution method. Of the remaining 187 susceptible isolates identified by mPCR, 183 isolates had MIC < or = 8 microg/ml (susceptible), 3 with MIC of 16 (intermediate resistant) and 1 with MIC of > or = 32 microg/ml (resistant). Comparing with the PCR-restriction enzyme analysis, mPCR concordantly identified 185 isolates either as being M. avium or M. intracellulare, whereas one isolate was misidentified and 32 isolates could not be identified. Comparing with reference methods, the mPCR showed the sensitivity, specificity, positive predictive and negative predictive value of 89, 100, 100, and 98% for detection of CLR resistance; 92, 98, 99, and 78% for identification of M. avium; and 57, 100, 100, and 89% for identification of M. intracellulare, respectively.

Francisella novicida bacteremia, Thailand.

We report isolation of Francisella novicida-causing bacteremia in a woman from Thailand who was receiving chemotherapy for ovarian cancer. The organism was isolated from blood cultures and identified by 16S rDNA and PPIase gene analyses. Diagnosis and treatment were delayed due to unawareness of the disease in this region.

Detection and differentiation between pathogenic and saprophytic Leptospira spp. by multiplex polymerase chain reaction.

A multiplex polymerase chain reaction (PCR) was developed for diagnosing leptospirosis and differentiating pathogenic and saprophytic leptospires. Specific primers were designed to amplify 23S rDNA from pathogenic Leptospira and saprophytic Leptospira spp. PCR products from 27 pathogenic and 5 (including 1 intermediate) saprophytic serovars were 615 and 316 base pairs (bp), respectively. After the restriction enzyme's digestion of PCR products, the fragments by SacI of pathogenic serovars and by PstI of saprophytic serovars were 339 and 276 bp and 202 and 114 bp, respectively. The PCR primers enabled amplification of DNA from L. meyeri serovar Ranarum as a pathogenic Leptospira spp. The PCR assay could detect 1 to 2 cells of leptospires and not amplify DNA from other 18 bacterial species. The sensitivity and specificity of this PCR in rat kidney, using isolation as gold standard, were 98.6% and 100%, respectively. The most appropriate sample preparation of blood for detecting DNA was buffy coat. Among the sample preparations from 7 laboratory-confirmed leptospirosis cases, leptospiral DNA was detected in all 7 buffy coat preparations, whereas leptospiral DNA was detected in only 3 plasma or serum samples. The PCR assay may be useful as a diagnostic tool for leptospirosis.

The efficacy of peer teaching for medical microbiology lectures.

This article was migrated. The article was marked as recommended. Background: A thorough understanding of infectious diseases is needed by medical professionals; therefore effective microbiological teaching is critical. Although faculty lectures are a convenient means of educating large groups of students, they may fail to engage students and convey an understanding of the subject. Therefore, we developed peer teaching methods based on game-based learning using a reality musical talent show format. Methods: A group of student representatives were trained to lecture to a class of 300 third-year medical students via a game show format over a 3-year period (2013-2015). Results: The students reported a higher level of understanding (3.6-4.2 vs 3.6-3.9 out of 5; p Conclusions: Peer teaching did improve the students' attitude towards learning and conferred teaching skills, but the learning activity needs adjustment to reduce the out-of-class preparation time.

Early-onset prosthetic valve endocarditis caused by Inquilinus sp.

Inquilinus is the newly described genus and has never been recognized as a cause of infective endocarditis. We report a case of early-onset prosthetic valve endocarditis caused by Inquilinus sp. in a tetralogy of Fallot patient who presented with heart failure. The bacterium was recovered from 11 consecutive blood cultures and identified by 16S rRNA gene sequencing.

Microbiology of primary acquired nasolacrimal duct obstruction: simple epiphora, acute dacryocystitis, and chronic dacryocystitis.

PURPOSE: The aim of this study was to determine the microbiology of primary acquired nasolacrimal duct obstruction (PANDO) and its antimicrobial susceptibilities. METHODS: Ninety-three patients (100 eyes) diagnosed with PANDO, categorized as acute, chronic dacryocystitis, or simple epiphora, were prospectively enrolled. Lacrimal sac contents were cultured for aerobic and anaerobic bacteria and fungi. Cultured organisms were identified, and antimicrobial susceptibility testing was performed for aerobic bacteria. RESULTS: Seventy-nine of the 100 samples were culture positive. One hundred twenty-seven organisms were isolated, and 29 different species were identified. Most microorganisms were Gram-positive bacteria (45 samples or 57.0% of all positive culture samples), whereas Gram-negative bacteria, anaerobic bacteria, and fungi were found in 39 (49.4%), 24 (30.4%), and four samples (5.1%), respectively. The most frequently isolated group was coagulase-negative staphylococci (27.8%), followed by nonspore-forming Gram-positive rods (anaerobe) (17.7%) and Pseudomonas aeruginosa (15.2%). Of the 100 samples, five, 45, and 50 samples were obtained from patients with acute dacryocystitis, chronic dacryocystitis, and simple epiphora, respectively. Subgroup analysis showed that Gram-negative organisms were isolated more frequently from the chronic dacryocystitis subgroup than from the simple epiphora subgroup (P=0.012). Antimicrobial susceptibility testing demonstrated that ciprofloxacin was the most effective drug against all Gram-positive and Gram-negative organisms. CONCLUSION: Patients with PANDO, with or without clinical signs of lacrimal infection, were culture positive. Gram-negative organisms were frequently isolated, which were different from previous studies. Ciprofloxacin was the most effective agent against all Gram-positive and Gram-negative organisms.

Sterility of non-preservative eye drops.

OBJECTIVES: To assess microbial contamination of common non-preservative eye drops stored at 4 degrees C and non-preservative fortified antibiotic eye drops used in a hospital inpatient setting. MATERIAL AND METHOD: A prospective study of the sterility of non-preservative eye drops was examined by dividing the patients into 2 groups. Group 1 composed of 5 samples of each ophthalmic preparation, which consisted of fortified antibiotics (vancomycin, cefazolin, gentamicin, and amikacin), non-preservative steroid eye drops (1% methylprednisolone and 0.01% dexamethasone), and 20% autologous serum stored at 4 degrees C. Group 2 was composed of 28 samples of fortified vancomycin and 23 samples of fortified amikacin used in an inpatient setting for 1 week and kept in 4 degrees C for 28 days. The contamination was assessed at the day of preparation and then at day 4, 7, 14, 21, 28 of all medications except autologous serum was done up to day 84. The sterility was examined by the culturing on tryptic soy broth and thioglycolate broth. RESULTS: In group 1 and 2, no contamination was detected in any samples within 4 weeks and up to 12 weeks for autologous serum. CONCLUSION: Non- preservative topical eye drops can be safely kept at 4 degrees C without contamination for 4 weeks and 12 weeks for 20% autologous serum drop. For topical antibiotics, under a strict protocol for preparation and usage might be used without significant risk of bacterial contamination for 4 weeks.

Pseudoclavibacter otitis media in a 3-year-old boy with pulmonary and spinal tuberculosis.

Pseudoclavibacter has rarely been documented as an etiologic agent of infection in humans. We presented the first case report of Pseudoclavibacter otitis media in a boy with pulmonary and spinal tuberculosis.A 3-year-old boy was referred to our hospital due to prolonged fever and progressive paraplegia for 3 months. He had yellowish discharge from both ear canals. The pleural fluid culture was positive for Mycobacterium tuberculosis. The discharge from both ears culture yielded yellow colonies of gram-positive bacilli with branching. This organism was positive for modified acid-fast bacilli stain but negative for acid-fast bacilli stain. Biochemical characteristics of this isolate were positive for catalase test but negative for oxidase, nitrate, esculin, and sugar utilization tests. The organism was further subjected to be identified by 16S ribosomal deoxyribonucleic acid gene sequencing. The result yielded Pseudoclavibacter species (99.4% identical), which could be most likely a potential pathogen in immunocompromised host like this patient. He responded well with intravenous trimetroprim-sulfamethoxazole for 6 weeks.This is the first case report of Pseudoclavibacter otitis media in children, and this case could emphasize Pseudoclavibacter species as a potential pathogen in immunocompromised host.

Nosocomial spread of the integron-located veb-1-like cassette encoding an extended-pectrum beta-lactamase in Pseudomonas aeruginosa in Thailand.

The beta-lactamase gene content and epidemiology of ceftazidime-resistant Pseudomonas aeruginosa isolates (24% of the total number of P. aeruginosa isolates) were investigated at a University Hospital in Thailand during a 4-month period in 1999. Of 33 nonrepetitive clinical isolates, 31 produced a VEB-1-like clavulanic acid-inhibited extended-spectrum beta-lactamase (ESBL). These isolates belonged to different pulsed-field gel electrophoresis types and subtypes. In 1 case, the bla(VEB-1)-like gene was plasmid located. The bla(VEB-1)-like genes were present as a gene cassette on class 1 integrons that varied in size and structure. In most cases, the veb-1 cassette was associated with an arr-2 cassette (rifampin resistance), aminoglycoside resistance gene cassettes, and an oxa-10-like cassette encoding a narrow-spectrum oxacillinase-type beta-lactamase. The present study indicates that ESBLs may be endemic in P. aeruginosa and illustrates that integrons are efficient means for their spread.

Shiga toxin- and enterotoxin-producing Escherichia coli isolated from subjects with bloody and nonbloody diarrhea in Bangkok, Thailand.

A total of 314 stool samples collected from 92 subjects with bloody diarrhea, 119 subjects with non-bloody diarrhea and 103 normal subjects in Bangkok, Thailand, were investigated for the presence of Shiga toxin-producing Escherichia coli (STEC) and enterotoxin-producing E. coli (ETEC) by multiplex PCR assay. Virulence genes and cytotoxic effect to Vero cells of STEC were also determined. STEC (5 isolates) and ETEC (18 isolates) were detected in 3 and 14 subjects, respectively. Among subjects containing ETEC, only one person belonged to normal control group. The detected STEC included two isolates (serotypes O26:H(-) and O111:H(-)) of Shiga toxin type 1 (Stx1-only) STEC from a child with non-bloody diarrhea, two isolates (Stx1-Stx2 STEC and Stx1-only STEC) from an adult with bloody diarrhea, and one isolate of Stx1-Stx2v STEC (O157:H7) from normal child. Only Stx1-Stx2 STEC isolate was found to exhibit toxicity to Vero cells and carry hlyA gene. The intimin encoding gene locus eaeA was not detected in any isolate. These results indicate that most of STEC isolates in Thailand were low virulent.

Puerperal retroperitoneal abscess caused by Clostridium difficile: case report and review of the literature.

BACKGROUND: Retroperitoneal infection can be lethal. Optimal management is still elusive to describe because of the small number of case reports. We presented here a case of retroperitoneal abscess caused by Clostridium difficile arising in the puerperal period. METHODS: Case report and review of recent English-language literature. RESULTS: The patient presented with surgical incision dehiscence. A gas-forming fluid collection was discovered in the pelvic retroperitoneal fascia by computed tomography, but the patient did not show marked symptoms of sepsis. Emergency laparotomy drainage and debridement were performed. Clostridium difficile was isolated, and she was treated with a three-week course of vancomycin. The patient recovered without major morbidity. Recent case reports describe variation in the course of the disease and management options for puerperal retroperitoneal infection. CONCLUSION: Puerperal retroperitoneal abscess caused by C. difficile can present with minimal symptoms. Prompt recognition, early surgical intervention, and optimal use of antibiotics can reduce morbidity and prevent death.

Post-partum, post-sterilization tubo-ovarian abscess caused by Fusobacterium necrophorum: a case report.

INTRODUCTION: Post-partum, post-sterilization tubo-ovarian abscess is a rare event. Fusobacterium necrophorum subspecies funduliforme, a normal flora found mainly in the oral cavity, appears to be the etiologic organism. CASE PRESENTATION: In this case report, a 25-year-old Thai woman had a post-partum, post-sterilization tubo-ovarian abscess caused by the strictly anaerobic bacterium, Fusobacterium necrophorum subspecies funduliforme. Progressively severe symptoms started 3 weeks after her third vaginal delivery with a tubal sterilization on the following day. On admission, she presented with peritonitis and impending shock. An exploratory laparotomy showed a ruptured left tubo-ovarian abscess. A segment of her ileum had to be resected because of severe inflammation. CONCLUSIONS: Fusobacterium necrophorum subspecies funduliforme can be an etiologic organism of a ruptured tubo-ovarian abscess following tubal sterilization in a healthy host.