bluuebunny/crossref_metadata_2025 huggingface dataset:
An outbreak of listeriosis suspected to have been caused by rainbow trout
Authors: Ericsson H , Eklöw A , Danielsson-Tham M L , Loncarevic S , Mentzing L O , Persson I , Unnerstad H , Tham W
Published: January 2020

An outbreak of listeriosis in Sweden, consisting of nine cases, was investigated by means of molecular typing of strains from patients and strains isolated from suspected foodstuffs, together with interviews of the patients. Listeria monocytogenes was isolated from six of the patients, and all isolates were of the same clonal type. This clonal type was also isolated from a "gravad" rainbow trout, made by producer Y, found in the refrigerator of one of the patients. Unopened packages obtained from producer Y were also found to contain the same clonal type of L. monocytogenes. Based on the interview results and the bacteriological typing, we suspect that at least six of the nine cases were caused by gravad or cold-smoked rainbow trout made by producer Y. To our knowledge, this is the first rainbow trout-borne outbreak of listeriosis ever reported.
Scytalidium dimidiatum and Lecythophora hoffmannii: unusual causes of fungal infections in a patient with AIDS
Authors: Marriott D J , Wong K H , Aznar E , Harkness J L , Cooper D A , Muir D
Published: January 2020

Immunocompromised patients are susceptible to infections by fungi that seldom cause disease in humans. We describe a human immunodeficiency virus-infected patient who had simultaneous infections with two fungi which are rare causes of serious infection: Lecythophora hoffmannii, causing chronic sinusitis, and Scytalidium dimidiatum, causing skin lesions, lymphangitis, and lymphadenitis. The clinical and pathologic findings are discussed.
Genes coding for enterotoxins and verotoxins in porcine Escherichia coli strains belonging to different O:K:H serotypes: relationship with toxic phenotypes
Authors: Blanco M , Blanco J E , Gonzalez E A , Mora A , Jansen W , Gomes T A , Zerbini L F , Yano T , de Castro A F , Blanco J
Published: January 2020

Seventy-four E. coli strains isolated from piglets with diarrhea or edema disease in Spain were serotyped and examined for production of heat-labile (LT) and heat-stable (ST) enterotoxins (LT-I, LT-II, STaH, STaP, and STb) and verotoxins (VT1, VT2, and VT2v = VTe) by phenotypic (Vero cell assay and infant mouse test) and genotypic (colony hybridization and PCR) methods. In general, an excellent correlation was found between the results obtained with a PCR approach and those determined with biological assays. DNA probes used in the hybridization also showed a very good agreement with phenotypic results, with the exception of a VT1 probe that initially produced 10 false-positive reactions. The gene coding for STb (58 strains) was the most prevalent gene detected by PCR, followed by those coding for STa (46 strains), LT (19 strains), VT2v (11 strains), and VT1 (1 strain). Apparently, in Spain three seropathotypes are predominant: (i) O149:K91:H10 K88+ LT-I+ STb+, (ii) O141:K85ab:H- P987+ STaP+, and (iii) O138:K81:H14 or H- STaP+ VT2v+. We conclude that PCR is a fast, specific, and practical method for the identification of enterotoxin and VT genes in clinical and epidemiological studies.
Fecal carriage of vancomycin-resistant enterococci in hospitalized patients and those living in the community in The Netherlands
Authors: Endtz H P , van den Braak N , van Belkum A , Kluytmans J A , Koeleman J G , Spanjaard L , Voss A , Weersink A J , Vandenbroucke-Grauls C M , Buiting A G , van Duin A , Verbrugh H A
Published: January 2020

In order to determine the prevalence of vancomycin-resistant enterococci (VRE) in The Netherlands, 624 hospitalized patients from intensive care units or hemato-oncology wards in nine hospitals and 200 patients living in the community were screened for VRE colonization. Enterococci were found in 49% of the hospitalized patients and in 80% of the patients living in the community. Of these strains, 43 and 32%, respectively, were Enterococcus faecium. VRE were isolated from 12 of 624 (2%) and 4 of 200 (2%) hospitalized patients and patients living in the community, respectively. PCR analysis of these 16 strains and 11 additional clinical VRE isolates from one of the participating hospitals revealed 24 vanA gene-containing, 1 vanB gene-containing, and 2 vanC1 gene-containing strains. All strains were cross-resistant to avoparcin but were sensitive to the novel glycopeptide antibiotic LY333328. Genotyping of the strains by arbitrarily primed PCR and pulsed-field gel electrophoresis revealed a high degree of genetic heterogeneity. This underscores a lack of hospital-driven endemicity of VRE clones. It is suggested that the VRE in hospitalized patients have originated from unknown sources in the community.
Analysis of an outbreak of non-phage-typeable methicillin-resistant Staphylococcus aureus by using a randomly amplified polymorphic DNA assay
Authors: Tambic A , Power E G , Talsania H , Anthony R M , French G L
Published: January 2020

A cluster of methicillin-resistant Staphylococcus aureus (MRSA) infections among patients on an intensive care unit (ICU) was detected by routine infection control surveillance. In the period from 5 January to 22 June 1995, 10 patients on the ICU and a further 6 patients (5 on one ward that had received colonized patients transferred from the ICU) were affected by MRSA strains with the same antibiotic susceptibility patterns. Seven (44%) of these 16 colonized patients developed MRSA bacteremia. MRSA isolates with the same characteristics were also found on the hands of one member of the ICU staff. The isolates were untypeable by phage typing, but 15 of 17 outbreak strains analyzed genetically had identical randomly amplified polymorphic DNA (RAPD) and pulsed-field gel electrophoresis (PFGE) profiles. A single strain of MRSA that was nontypeable by phage typing and that was isolated on the ICU on 1 January and six nontypeable and epidemiologically unrelated MRSA isolates all had RAPD profiles distinct from that of the outbreak strain. Implementation of strict infection control measures stopped the further spread of MRSA on the ICU, the affected general ward, and seven other wards that received MRSA carriers from the ICU. Although nontypeable by phage typing and not previously recognized as an epidemic strain, this strain of MRSA was readily transmissible and highly virulent. RAPD typing was found to be a simple, rapid, and effective method for the epidemiological investigation of this outbreak, and performance of typing by this method was simpler and less time-consuming than that of typing by PFGE. RAPD typing may have more general application for the study of S. aureus infections in hospitals.
Human papillomavirus and host variables as predictors of clinical course in patients with juvenile-onset recurrent respiratory papillomatosis
Authors: Gabbott M , Cossart Y E , Kan A , Konopka M , Chan R , Rose B R
Published: January 2020

This study provides the first systematic evaluation of papillomavirus type and viral mutation occurring during the course of juvenile-onset recurrent respiratory papillomatosis. One hundred ninety-nine consecutive papillomas excised from 47 children between 1981 and 1996 at The New Children's Hospital in Sydney, Australia, were tested for human papillomavirus (HPV) DNA by PCR. PCR products from the viral upstream regulatory region (URR) enhancer were sequenced, and variation was related to clinical variables. Forty-four of the 47 children had HPV-induced papillomas, with type 11 accounting for 24 (55%) and type 6 accounting for 19 (43%); one (2%) was positive for either type 6 or 11. Overall, 183 (98%) of the 186 samples with amplifiable DNA were HPV positive. There was no change in HPV type over time and no statistically significant association between HPV type and disease aggressiveness. One novel, large-scale URR duplication was identified in an HPV type 11 isolate in the last of a series of six papillomas examined and the first from the bronchus. However, the duplication was not found in HPV type 11 isolates from the associated pulmonary carcinoma and its metastases in other organs. Three of 14 URR point mutations coincided with transcription factor binding sites, but there were no obvious associations with clinical course. Chi-square and multiple linear regression analyses of clinicopathological variables revealed early age at diagnosis (less than 4 years) as an independent predictor of aggressive disease (P < 0.001). A bimodal distribution of the age at diagnosis was noted, with peaks at 2 and 11 years of age.
Evaluation of six commercial kits for detection of human immunoglobulin M antibodies to Toxoplasma gondii. The FDA Toxoplasmosis Ad Hoc Working Group
Authors: Wilson M , Remington J S , Clavet C , Varney G , Press C , Ware D
Published: January 2020

As a result of reports received by the Food and Drug Administration (FDA) of false-positive results obtained with FDA-cleared in vitro diagnostic kits for the detection of Toxoplasma-specific human immunoglobulin M (IgM) antibodies, an FDA-sponsored evaluation of six kits was performed. A battery of 258 serum specimens, including 30 specimens drawn 1 to 5 months after initial Toxoplasma infection and 228 specimens from Toxoplasma IgG-positive individuals, Toxoplasma IgG-negative individuals, rheumatoid factor-positive persons, and persons determined to be Toxoplasma IgM positive by commercially available assays, was assembled, randomly assorted, and coded. The battery was tested at the FDA with six commercially available kits, at the Palo Alto Medical Foundation (PAMF) by the PAMF double-sandwich IgM enzyme-linked immunosorbent assay (PAMF IgM ELISA), and at the Centers for Disease Control and Prevention (CDC) by the CDC EIA IgM. The results of the PAMF IgM ELISA that were obtained with the battery were considered to be the "gold standard" for this study; specificity rates were computed by considering the PAMF results to be 100% specific. Sensitivity and specificity rates were found to be as follows: CDC EIA IgM, 100 and 99.1%, respectively; Abbott IMx Toxo IgM, version 1, 100 and 77.5%, respectively; Abbott IMx Toxo IgM, version 2, 93.3 and 97.3%, respectively; Abbott Toxo-M EIA, 100 and 84.2%, respectively; BioMérieux Vitek VIDAS Toxo IgM, 100 and 98.6%, respectively; BioWhittaker Toxocap-M, 100 and 95.9%, respectively; Gull Toxo IgM, 97 and 85.6%, respectively; and Sanofi Diagnostics Pasteur Platelia Toxo IgM, 100 and 96.8%, respectively. Although the extent of false-positive reactions with these kits cannot be calculated because the study was retrospective and sample choices were biased, the results may be useful as an indicator of the relative specificities of these kits.
Evaluation of the MB/BacT system and comparison to the BACTEC 460 system and solid media for isolation of mycobacteria from clinical specimens
Authors: Rohner P , Ninet B , Metral C , Emler S , Auckenthaler R
Published: January 2020

The MB/BacT automated system is designed for the isolation of mycobacteria from clinical specimens. It utilizes a colorimetric sensor and reflected light to continuously monitor the CO2 concentration in the culture medium. We compared its performance to that of the BACTEC 12B media for the radiometric BACTEC 460 instrument and that of solid culture media. Respiratory specimens and urine samples were decontaminated with 4% NaOH. The vials of the two instruments were inoculated with 500 microl of sample and two solid egg-based media at 200 microl each. All vials were incubated at 37 degrees C for 6 weeks. A total of 1,078 specimens (633 respiratory specimens, 78 cerebrospinal fluid specimens, 177 other body fluid specimens, 87 urine specimens, and 103 other types of specimens) were cultured in parallel. Mycobacteria could be identified from 73 (6.8%) specimens: 67 M. tuberculosis, 3 M. kansasii, 1 M. xenopi, 1 M. terrae, and 1 mixed M. avium with M. scrofulaceum. Of these, 63 (86.3%) specimens were positive with the MB/BacT system, 67 (91.8%) were positive with the BACTEC 460 instrument, and 58 (79.5%) were positive with the two egg-based media. MB/BacT cultures were positive on average after 17.5 (+/-6.4) days, BACTEC cultures with a growth index of >20 (mean, 200) were positive after 14.3 (+/-8.2) days, and egg-based media were positive after 24.2 (+/-7.5) days. Microorganisms other than mycobacteria contaminated 46 (4.3%) MB/BacT cultures and 31 (2.9%) BACTEC cultures, which had to be discarded. The MB/BacT system is a well-automated system for the detection of M. tuberculosis in clinical specimens without using radioactive reagents. Further trials are required to determine whether it is suitable for the culture of nontuberculous mycobacteria.
Randomly amplified polymorphic DNA PCR for comparison of Mycobacterium abscessus strains from nosocomial outbreaks
Authors: Zhang Y , Rajagopalan M , Brown B A , Wallace R J
Published: January 2020

Mycobacterium abscessus is an important cause of water-related nosocomial outbreaks or pseudo-outbreaks. Strain comparison has relied on pulsed-field gel electrophoresis (PFGE). Unfortunately, almost 50% of strains cannot be assessed by this method. We studied 118 strains of M. abscessus previously studied by PFGE by randomly amplified polymorphic DNA (RAPD) PCR, including isolates from eight nosocomial outbreaks. Ten random primers were evaluated by using DNA prepared by boiling or phenol-chloroform extraction. Both DNA preparations gave the same grouping of isolates for three outbreaks compared to the groupings obtained by PFGE. Five outbreaks due to M. abscessus which gave broken DNA by PFGE gave evaluable patterns when studied by RAPD-PCR, with isolate clustering being consistent with that from other laboratory and epidemiologic data. The patterns were highly method dependent, strain comparison required the use of multiple primers, and the method worked best with purified DNA and by using strains for comparison on the same gel. We propose categories of indistinguishable, different, and inconclusive when comparing strains by RAPD-PCR. This study demonstrates that RAPD-PCR can be used for genetic comparison of M. abscessus strains, including strains which cannot be compared by PFGE, but the potential for misinterpretation is greater than that by PFGE.
Reversed passive latex agglutination assay for detection of toxigenic Corynebacterium diphtheriae
Authors: Toma C , Sisavath L , Iwanaga M
Published: January 2020

A reversed passive latex agglutination (RPLA) assay for determining the toxigenicity of Corynebacterium diphtheriae is presented. Rabbit antitoxin antiserum was raised by using commercially available diphtheria toxoid. This antiserum reacted with the diphtheria toxin when the culture supernatant was assayed by Western blotting, and it did not cross-react with other extracellular antigens. Affinity-purified antibodies for latex sensitization were obtained by using a Hi Trap N-hydroxysuccinimide-activated column. Demonstration of toxin in five of seven clinical isolates was in accordance with the PCR assay and the Vero cell cytotoxicity test. Culture of the bacteria for 6 h was sufficient for toxin production, and an additional 6 h was needed to observe latex agglutination. Therefore, diphtheria toxin can be detected in 12 h by this method. The lowest concentration of diphtheria toxin detectable by the RPLA assay was about 5 ng/ml. The RPLA assay can provide a convenient and reliable method for laboratories involved in the identification of toxinogenic corynebacteria.
Clinical and epidemiological significance of enterococci intrinsically resistant to vancomycin (possessing the vanC genotype)
Authors: Toye B , Shymanski J , Bobrowska M , Woods W , Ramotar K
Published: January 2020

Constitutive low-level vancomycin resistance is found intrinsically in certain enterococcal species and is encoded by vanC ligase genes. These intrinsically vancomycin-resistant enterococci (VRE) will be referred to as VANC VRE. A prospective study to determine the clinical and epidemiologic significance of VANC VRE was conducted. VANC VRE were recovered from the stools of 34 of 601 (5.7%) patients, a rate similar to that obtained for the stools of 100 outpatients in the community (5%). VANC VRE were also isolated from the nonstool specimens of 9 of 538 patients (1.7%), including two patients with bacteremia. No VRE of the vanA or vanB genotypes were detected in nonstool specimens. Eighty-two hospital contacts of the first 23 patients found to be colonized or infected with VANC VRE were screened, and 6 contacts were found to be gastrointestinal carriers of VANC VRE. However, typing of isolates from these 6 contacts by pulsed-field gel electrophoresis with SmaI showed the isolates to be unique and different from those recovered from the index patients. In fact, all VANC VRE isolates from different patients in this study were unique. A case-control study with patients who were negative when screened for VANC VRE as controls failed to identify any risk factor associated with colonization or infection with this organism. VANC VRE were infrequently recovered from clinical specimens but were occasionally found as part of the normal stool flora. Since no transmission between patients was documented, additional isolation procedures may not be necessary for patients colonized or infected with VANC VRE.
Comparison of use of phenotypic and genotypic characteristics for identification of species of the anamorph genus Candida and related teleomorph yeast species
Authors: Latouche G N , Daniel H M , Lee O C , Mitchell T G , Sorrell T C , Meyer W
Published: January 2020

A total of 49 type and neotype isolates and 32 clinical isolates of the anamorph genus Candida and related teleomorph genera were obtained from different culture collections and clinical laboratories. Isolates were subjected to two phenotypic methods of identification, Vitek yeast biochemical card (YBC) and API ID 32C, both based on carbohydrate assimilation, and one genotypic method, PCR fingerprinting, based on the detection of DNA polymorphisms between minisatellite-specific sequences with the primer M13 (5' GAGGGTGGCGGTTCT 3'). The correct identification of a strain at the Centraalbureau voor Schimmelcultures was used as the gold standard for the identification of an isolate. When the study was restricted to species included in the respective biochemical databases, the Vitek YBC and API ID 32C systems performed adequately with positive identification rates of 87.3 and 76.8%, respectively. When uncommon species were added to the study, several of which are not included in the databases, the identification efficiencies were 76.5 and 77.5%, respectively. By comparison, all isolates were correctly identified by PCR fingerprinting, with 63 reference species profiles in the databank. Sufficient polymorphisms among the total set of banding patterns were observed, with adequate similarity in the major patterns obtained from a given species, to allow each isolate to be assigned unambiguously to a particular species. In addition, variations in minor bands allowed for differentiation to the strain level. PCR fingerprinting was found to be rapid, reproducible, and more cost-effective than either biochemical approach. Our results provide reference laboratories with an improved identification method for yeasts based on genotypic rather than phenotypic markers.
Fecal microflora in a patient with short-bowel syndrome and identification of dominant lactobacilli
Authors: Kaneko T , Bando Y , Kurihara H , Satomi K , Nonoyama K , Matsuura N
Published: January 2020

Fecal microflora and lactate concentrations in blood and feces obtained from a patient (a 5 year-old boy) with short-bowel syndrome (SBS) were compared during acidosis to results for the normal condition (no SBS symptoms). The taxonomical position of the lactobacilli found predominantly in the feces sample obtained 2 days before the fifth attack was also studied. The D-lactate level in serum obtained 1 day after the fourth attack was 10-fold higher than that for the normal condition, although there was not a great difference in L-lactate levels. D-Lactate (3.91 mM) and L-lactate (2.86 mM) were also detected in the feces samples collected 2 days before the fifth attack, while no lactate was detected in the feces sample for the normal condition. The counts of total fecal bacteria, especially anaerobic bacteria such as members of the family Bacteroidaceae, were found to be low. The counts of lactobacilli and the total population of lactobacilli relative to total fecal bacteria in the feces 2 days before the fifth attack (40.4%) were extremely high. In this case, a majority of the lactobacilli were D-lactate producers as determined by homolactic fermentation. These lactobacilli were identified as Lactobacillus delbrueckii subsp. lactis. The percentages of bifidobacteria relative to total fecal bacteria in feces samples obtained both 2 days before the fifth attack (50.9%) and for normal condition (61.9%) were also high, although these bacteria were L-lactate producers. In the feces samples for the normal condition, the D-lactate producers decreased to less than 10(9) per g, while the counts of L- or DL-lactate producers were 100-fold higher than the numbers in feces samples obtained 2 days before the fifth attack. These results suggested that an increase in the level of D-lactate producers, such as L. delbrueckii subsp. lactis, in the colon may be associated with the clinical expression of metabolic acidosis.
Human granulocytic ehrlichiosis in southern Germany: increased seroprevalence in high-risk groups
Authors: Fingerle V , Goodman J L , Johnson R C , Kurtti T J , Munderloh U G , Wilske B
Published: January 2020

To date, human granulocytic ehrlichiosis (HGE), the causative agent of which is likely transmitted by ticks in the Ixodes ricinus-Ixodes persulcatus complex, has not been diagnosed with certainty in patients outside the United States. The presence of a closely related vector tick, I. ricinus, as well as the occurrence of similar Ehrlichia spp. of veterinary importance, suggests that this disease is likely to be present in Europe. The aim of the present study was to compare the prevalence of antibodies against the HGE agent in sera collected from patients in groups at high risk for exposure to I. ricinus with that of a control population. Risk groups consisted of 150 forestry workers and 105 patients with an established diagnosis of Lyme disease. The control group was 103 healthy blood donors without a history of tick bites. We used a patient isolate of the HGE agent from Minnesota (J. L. Goodman, C. Nelson, B. Vitale, J. E. Madigan, J. S. Dumler, T. J. Kurtti, and U. G. Munderloh, N. Engl. J. Med. 334:209-215, 1996) propagated in HL60 cells as the source of antigen for a specific immunofluorescence assay (IFA). Elevated IFA titers (> or = 1:80) were present in 21 of 150 (14%) serum samples from forestry workers and in 12 of 105 (11.4%) serum samples from Lyme disease patients, but in only 2 of 103 (1.9%) serum samples from blood donors (P < or = 0.01 for either of the at-risk groups versus blood donors). The results of this study suggest that the HGE agent or a closely related organism exists in southern Germany and that seroconversion to it is common among groups exposed to Ixodes ticks. Final proof that HGE occurs in Germany will require the isolation of the causative agent from patients. HGE should be considered in the differential diagnosis of febrile illnesses in individuals exposed to Ixodes ticks in Europe as well as in North America.
Incidence and risk factors for hepatitis C among injection drug users in Baltimore, Maryland
Authors: Villano S A , Vlahov D , Nelson K E , Lyles C M , Cohn S , Thomas D L
Published: January 2020

Between 1988 and 1996, the incidence of and risk factors for hepatitis C virus (HCV) infection were studied in a cohort of injection drug users in Baltimore, Maryland. By second-generation antibody testing of stored serum samples, 142 participants were found to be susceptible to HCV at the time they entered the study. After a median follow-up of 6.5 years, 43 participants (30.3%) developed antibodies to HCV (anti-HCV). The overall incidence was 6.4 cases per 100 person-years, but a substantial decline in the annual incidence rate was observed after the first 2 years (1988 to 1990, 13.4/100 person-years; 1991 to 1996, 2.3/100 person-years [P = 0.0001 for trend]). Participants who acknowledged active drug use, especially those who acknowledged frequent use and sharing of drug paraphernalia, were at increased risk of HCV infection. However, high-risk sexual practices were not associated with HCV seroconversion. Efforts to reduce HCV infection must be focused on curbing drug use and especially on the sharing of needles and drug paraphernalia.
Citrobacter farmeri bacteremia in a child with short-bowel syndrome
Authors: Bruckner D A , Colonna P , Glenn D , Abbott S L , Janda J M
Published: January 2020

A case of sepsis in a pediatric patient due to the newly described Citrobacter species C. farmeri is described. Factors predisposing this child to infection included short-bowel syndrome requiring total parenteral nutrition.
Salivary immunoglobulin G assay to diagnose Helicobacter pylori infection in children
Authors: Luzza F , Oderda G , Maletta M , Imeneo M , Mesuraca L , Chioboli E , Lerro P , Guandalini S , Pallone F
Published: January 2020

An in-house enzyme-linked immunosorbent assay (ELISA) for measurement of Helicobacter pylori-specific immunoglobulin G (IgG) and IgA in saliva was evaluated by comparison with histopathologic (Giemsa staining) and biochemical (urease quick test) examination of gastric biopsy specimens obtained from 112 children referred for diagnostic gastroscopy. Serum H. pylori IgG was also measured in a subgroup of 50 children by the same ELISA. Salivary H. pylori IgG levels were significantly higher in H. pylori-positive (n = 57) than in H. pylori-negative (n = 55) children (P < 0.001). The sensitivity and specificity of the salivary IgG test were 93 and 82%, respectively; the positive and negative predictive values were 84 and 92%, respectively; and the accuracy was 87.5%. Salivary H. pylori IgA did not distinguish H. pylori-positive from H. pylori-negative children. The performance of serum H. pylori IgG was slightly (3 to 6%) better than that of salivary H. pylori IgG. The salivary IgG test can be considered a useful tool for the screening of H. pylori infection in children.
Identification of Serpulina species associated with porcine colitis by biochemical analysis and PCR
Authors: Fellström C , Pettersson B , Thomson J , Gunnarsson A , Persson M , Johansson K E
Published: January 2020

A PCR system for the detection and identification of group IV spirochetes (Serpulina pilosicoli) was designed to complement biochemical tests, e.g., the hippurate hydrolysis and beta-glucosidase tests, and to verify the accuracy of a previously proposed biochemical classification system. The PCR assay was based on amplification of a segment of the 16S rRNA gene. Both primers were constructed to selectively amplify the 16S rRNA gene of Serpulina pilosicoli. All analyzed Serpulina strains exhibiting the capacity to hydrolyze hippurate and lacking beta-glucosidase activity, including the type strain for spirochetal diarrhea, P43, were amplified with the PCR system. All other tested strains, including type and field strains of different phenotypes of Serpulina species, as well as Salmonella species, Campylobacter species, and Escherichia coli strains, were negative in the assay. Among the tested strains were 18 Scottish field isolates originating from the mucosae of pigs with colitis. A simple classification scheme, suitable for routine classification of porcine intestinal spirochetes, is also presented. The scheme is based on hemolysis, indole production, and the hippurate hydrolysis test.
Case report: field-acquired subclinical Babesia equi infection confirmed by in vitro culture
Authors: Holman P J , Hietala S K , Kayashima L R , Olson D , Waghela S D , Wagner G G
Published: January 2020

A horse with no prior clinical history of equine piroplasmosis tested negative for Babesia caballi and Babesia equi in the complement fixation test before importation into the United States from France. After 5 years in residence in the United States, the animal tested serologically positive for B. equi by the complement fixation test, the immunofluorescent antibody test, and Western blot analysis. The carrier status of the horse was confirmed by culture of B. equi parasites. In vitro culture offers an efficient and comparatively inexpensive method to determine the carrier status of horses suspected of harboring B. equi.
Bacteremia caused by Arcobacter cryaerophilus 1B
Authors: Hsueh P R , Teng L J , Yang P C , Wang S K , Chang S C , Ho S W , Hsieh W C , Luh K T
Published: January 2020

Arcobacter cryaerophilus group 1B, a gram-negative, curved or helical bacillus primarily known as a bovine and porcine pathogen, was isolated from the blood of a uremic patient with hematogenous pneumonia. The patient was treated successfully with ceftizoxime and tobramycin.
Study of an outbreak of cefoxitin-resistant Klebsiella pneumoniae in a general hospital
Authors: Gazouli M , Kaufmann M E , Tzelepi E , Dimopoulou H , Paniara O , Tzouvelekis L S
Published: January 2020

During a 3-month period, six Klebsiella pneumoniae isolates resistant to cefoxitin and penicillin-inhibitor combinations were derived from patients in the intensive care unit of a hospital in Athens, Greece. Enterobacterial repetitive intergenic consensus PCR and pulsed-field gel electrophoresis provided evidence of the clonal origin of the isolates. Conventional techniques and ribotyping were inadequate in proving that the isolates were related. Resistance was due to a plasmidic class C beta-lactamase.
Comparison of PCR and hybrid capture methods for detection of human papillomavirus in injection drug-using women at high risk of human immunodeficiency virus infection
Authors: Shah K V , Solomon L , Daniel R , Cohn S , Vlahov D
Published: January 2020

We compared Hybrid Capture, a new technique for detection of human papillomaviruses (HPV), with a PCR assay based on L1 consensus primers. By both methods, the HPV prevalence was higher in human immunodeficiency virus (HIV)-positive women than in HIV-negative women. PCR had a higher sensitivity (0.89 versus 0.48) but lower specificity (0.43 versus 0.93) for detection of Pap smear abnormalities, compared to Hybrid Capture. The higher intensity of hybridization signal by PCR was related to higher estimates of viral load by Hybrid Capture.
Quantitation of cytomegalovirus (CMV) DNA in leukocytes of human immunodeficiency virus-infected subjects with and without CMV disease by using PCR and the SHARP Signal Detection System
Authors: Boivin G , Handfield J , Murray G , Toma E , Lalonde R , Lazar J G , Bergeron M G
Published: January 2020

We report the development of a simple and rapid PCR assay for quantitation of the cytomegalovirus (CMV) DNA load in polymorphonuclear leukocytes. Using this system, a very good correlation was found between a high number of CMV copies in the blood and the presence of CMV disease in subjects with AIDS.
PCR identification of Mycobacterium bovis BCG
Authors: Talbot E A , Williams D L , Frothingham R
Published: January 2020

The attenuated bacillus Calmette-Guérin (BCG) vaccine strain is derived from a virulent strain of Mycobacterium bovis. BCG is difficult to differentiate from other strains of M. bovis and other members of the M. tuberculosis complex by conventional methods. Recently, a genomic region designated RD1 was found to be present in all virulent M. bovis and M. tuberculosis strains tested but deleted from all BCG strains tested. With this information, a multiplex PCR method was developed to detect the RD1 deletion. A large collection of BCG and other M. tuberculosis complex strains from diverse host and geographic origins was tested. RD1 was deleted in 23 of 23 BCG strains. RD1 was present in 129 of 129 other M. tuberculosis complex strains. This multiplex PCR method can be used as a tool for the rapid and specific identification of BCG.
Diagnosis of triple-lumen catheter infection: comparison of roll plate, sonication, and flushing methodologies
Authors: Sherertz R J , Heard S O , Raad I I
Published: January 2020

In a recent clinical trial, 248 triple-lumen catheters were removed from patients in an intensive care unit, and their tip and subcutaneous segments were cultured by both the sonication and roll plate methods; for 191 of these catheters, flush cultures of all three catheter lumens were also performed. Previously published quantitative endpoints were used to define significant catheter colonization. By using a composite index as a definition of colonization (any of the seven types of cultures meeting quantitative criteria), sonication of the subcutaneous segment was the most sensitive at detecting colonization (58%), followed by sonication of the catheter tip (53%). Sonication of both the subcutaneous and tip segments was 20% more sensitive than sonication of an adjacent catheter segment by the roll plate method (P < 0.05). The greater sensitivity of the sonication method could be attributed to its greater ability than the roll plate method to detect catheter lumen colonization (82 versus 57%, respectively; P = 0.01). A greater number of positive catheter segment cultures were found for colonized catheters from patients with associated bacteremia than for colonized catheters from patients without bacteremia (57 versus 37%; P = 0.004), making any culture method more likely to identify them. For catheters with significant colonization of only one site, the localization was as follows: 36.7% subcutaneous segment, 36.7% catheter lumen, and 26.6% tip segment. These findings suggest that the current practice of culturing a single segment of a central vascular catheter is inadequate and needs to be reexamined. They further suggest that initial colonization of the catheter lumen and tip segments may be more important than previously thought and may require a change in thinking of strategies designed to prevent catheter infection.
Production and characterization of monoclonal antibodies specific for the lipopolysaccharide of Escherichia coli O157
Authors: Westerman R B , He Y , Keen J E , Littledike E T , Kwang J
Published: January 2020

Identification of the O157 antigen is an essential part of the detection of Escherichia coli O157:H7, which is recognized as a major etiologic agent of hemorrhagic colitis. However, polyclonal antibodies produced against E. coli O157:H7 lipopolysaccharide (LPS) may react with several other bacteria including Brucella abortus, Brucella melitensis, Yersinia enterocolitica O9, Escherichia hermannii, and Stenotrophomonas maltophilia. We produced eight monoclonal antibodies (MAbs) specific for the LPS of E. coli O157. Western blots (immunoblots) of both the phenol phase (smooth) and the aqueous phase (rough) of hot phenol-water-purified LPS indicated that three of the MAbs were specific for the O antigen and five were reactive with the LPS core. The eight MAbs could be further differentiated by their reactivities to Salmonella O30 LPS (group N), which is reported to be identical to the E. coli O157 antigen. All eight MAbs reacted strongly to all of the 64 strains of E. coli O157 tested, which included 47 isolates of O157:H7 and 17 other O157 strains. None of the eight MAbs cross-reacted with any of the 38 other E. coli serotypes tested, which consisted of 29 different O-antigen serotypes, or with 38 strains (22 genera) of non-E. coli gram-negative enteric bacteria.
A nosocomial outbreak due to Enterobacter cloacae strains with the E. hormaechei genotype in patients treated with fluoroquinolones
Authors: Davin-Regli A , Bosi C , Charrel R , Ageron E , Papazian L , Grimont P A , Cremieux A , Bollet C
Published: January 2020

During a 7-month period, we isolated 21 highly fluoroquinolone-resistant Enterobacter cloaecae strains in units from two hospitals in Marseille, France. Random amplification of polymorphic DNA showed clonal identity between isolates which, furthermore, presented the Enterobacter hormaechei genotype on DNA-DNA hybridization. The emergence of this clone was observed only in patients treated with fluoroquinolones.
The diagnostic yield of acid-fast-bacillus smear-positive sputum specimens
Authors: Stone B L , Burman W J , Hildred M V , Jarboe E A , Reves R R , Wilson M L
Published: January 2020

The yield of mycobacterial culture from acid-fast-bacillus smear-positive sputum specimens was 387 or 439 (88.2%). Forty-nine of 52 culture-negative specimens came from patients on treatment. We conclude that the yield of culture from smear-positive sputum specimens is very high and that only two acid-fast-bacillus smear-positive specimens are needed for the initial evaluation of pulmonary mycobacteriosis.
Species differentiation by internally transcribed spacer PCR and HhaI digestion of fluconazole-resistant Candida krusei, Candida inconspicua, and Candida norvegensis strains
Authors: Nho S , Anderson M J , Moore C B , Denning D W
Published: January 2020

PCR amplification of the regions containing the internally transcribed spacers and 5.8S rRNA gene of Candida krusei, C. inconspicua, and C. norvegensis yielded fragments of 510, 460, and 500 bp, respectively. HhaI digestion of these fragments yielded species-specific bands. Random amplification of polymorphic DNA with primer R108 showed interspecific discriminatory band patterns. Susceptibilities to fluconazole and amphotericin B were determined.
Improved detection of bacterial growth in continuous ambulatory peritoneal dialysis effluent by use of BacT/Alert FAN bottles
Authors: Alfa M J , Degagne P , Olson N , Harding G K
Published: January 2020

Culture-negative peritonitis is a major complication for patients on continuous ambulatory peritoneal dialysis (CAPD) and precludes organism-specific therapy. The aim of the present study was to compare inoculation of 10 ml of CAPD effluent into BacT/Alert blood culture bottles (FAN [fastidious antimicrobic neutralizing], BacTAlert aerobic [BTA], and BacT/Alert anaerobic [BTAn] bottles) to our conventional method of using 50 ml of concentrated CAPD effluent to inoculate peptone broth bottles (BD bottles) and MacConkey agar and blood agar medium (BA-MAC). The FAN, BTA, and BTAn bottles were monitored automatically in the BacT/Alert blood culture instrument. A total of 207 CAPD effluents were studied, and in 97 bacteria were detected by at least one method. Compared to BTA bottles (79 of 97; 81.4%), BTAn bottles (78 of 97; 80.4%), and BD bottles (88 of 97; 90.7%), the single best broth medium for detecting bacterial growth in CAPD effluents was the FAN bottle (90 of 97 effluents; 92.8%). A total of 125 bacterial species were detected by any method, and the majority (91.8%) of CAPD effluents were infected with a single species. A combination of FAN and BTAn bottles detected 111 of 125 (88.8%) of all organisms, whereas a combination of BD bottles and BA-MAC detected 107 of 125 (85.6%) of all organisms. One or more organisms that would have been completely missed by the conventional method with BD bottles and BA-MAC were detected in 18 CAPD effluents. Of these 18 CAPD effluents, 6 showed no growth by the conventional method with BD bottles and BA-MAC. On the basis of our data, the most sensitive and least labor intensive method was direct inoculation of 10 ml of CAPD effluent into a FAN bottle and a BTAn bottle, which could be automatically monitored by the BacT/Alert blood culture instrument. On the basis of case definitions for peritonitis, the sensitivities and specificities of the methods with FAN and BTAn bottles and with BD bottles and BA-MAC were 81.1 and 98.8% and 74.5 and 96.5%, respectively.
Comparison and application of ribosome spacer DNA amplicon polymorphisms and pulsed-field gel electrophoresis for differentiation of methicillin-resistant Staphylococcus aureus strains
Authors: Kumari D N , Keer V , Hawkey P M , Parnell P , Joseph N , Richardson J F , Cookson B
Published: January 2020

Analysis of sequences in the fragments of the 16S-23S rRNA intergenic spacer region by the ribosome spacer PCR (RS-PCR) can differentiate strains of methicillin-resistant Staphylococcus aureus (MRSA). We compared this technique with pulsed-field gel electrophoresis (PFGE) for typing MRSA strains and its application during an investigation of an outbreak. A total of 180 isolates of MRSA collected from various hospital laboratories within the United Kingdom and elsewhere were typed by PFGE and RS-PCR. PFGE identified 17 different types among the 180 strains examined, and RS-PCR generated 13 different types. PFGE could detect minor genetic variations among the isolates and could identify the variants which were not discriminated by RS-PCR. Four unique strain types detected by PFGE were not detected by RS-PCR. When applied to typing the outbreak-related strains from the vascular surgery unit at the General Infirmary at Leeds, the results of RS-PCR were identical to those of PFGE. Our results have shown that RS-PCR is a rapid, inexpensive technique that is highly reproducible and almost as discriminatory as PFGE for typing MRSA isolates and should be useful in the local investigation of MRSA outbreaks.
A sensitive, type-specific, fluorogenic probe assay for detection of human papillomavirus DNA
Authors: Swan D C , Tucker R A , Holloway B P , Icenogle J P
Published: January 2020

A simple method for the detection of a number of human papillomavirus (HPV) genotypes associated with cervical cancer has been developed. The assay exploits the 5'-->3' exonucleolytic activity of Taq DNA polymerase to increase the signal from fluorescent dyes by releasing them from genotype-specific probes during PCR. The probes are oligonucleotides with a 5' reporter dye (6-carboxyfluorescein), a quencher dye (6-carboxy-tetramethyl-rhodamine), and a phosphate-blocked 3' end. In the intact probe, the proximity of the reporter and the quencher results in suppression of reporter fluorescence by Förster-type energy transfer (V. T. Förster. Ann. Phys. 2:55-75, 1948). If the probe is bound downstream of either primer during PCR, the 5'-->3' exonucleolytic activity of Taq polymerase degrades it, allowing the reporter to diffuse away from the quencher, which results in an increase in reporter fluorescence. The increased fluorescence is directly related to the amount of target DNA and can be detected with an automated fluorometer. Probes for the L1 region of the cervical-cancer-associated HPV types 16, 18, 31, 33, and 35 were synthesized and the assays were optimized. The most sensitive assay can detect as few as two copies of HPV DNA in human cervical specimens.
Characterization of Escherichia coli O157:H7 and other Shiga toxin-producing E. coli serotypes isolated from sheep
Authors: Kudva I T , Hatfield P G , Hovde C J
Published: January 2020

The isolation and characterization of Escherichia coli O157:H7 and non-O157 Shiga toxin-producing E. coli (STEC) strains from sheep are described. One flock was investigated for E. coli O157:H7 over a 16-month period that spanned two summer and two autumn seasons. Variation in the occurrence of E. coli O157:H7-positive sheep was observed, with animals being culture positive only in the summer months but not in the spring, autumn, or winter. E. coli O157:H7 isolates were distinguished by pulsed-field gel electrophoresis (PFGE) of chromosomal DNA and toxin gene restriction fragment length polymorphism (RFLP) analysis. Ten PFGE patterns and five RFLP patterns, identified among the isolates, showed that multiple E. coli O157:H7 strains were isolated from one flock, that a single animal simultaneously shed multiple E. coli O157:H7 strains, and that the strains shed by individuals changed over time. E. coli O157:H7 was isolated only by selective enrichment culture off 10 g of ovine feces. In contrast, strains of eight STEC serotypes other than O157:H7 were cultured from feces of sheep from a separate flock without enrichment. The predominant non-O157 STEC serotype found was O91:NM (NM indicates nonmotile), and others included O128:NM, O88:NM, O6:H49, and O5:NM. Irrespective of serotype, 98% of the ovine STEC isolates possessed various combinations of the virulence-associated genes for Shiga toxin(s) and the attaching-and-effacing lesion (stx1, stx2, and eae), suggesting their potential for human pathogenicity. The most common toxin-eae genotype was positive for stx1, stx2, and eae. A Vero cell cytotoxicity assay demonstrated that 90% of the representative STEC isolates tested expressed the toxin gene. The report demonstrates that sheep transiently shed a variety of STEC strains, including E. coli O157:H7, that have potential as human pathogens.
Serologic and molecular detection of granulocytic ehrlichiosis in Rhode Island
Authors: Yeh M T , Mather T N , Coughlin R T , Gingrich-Baker C , Sumner J W , Massung R F
Published: January 2020

A new indirect fluorescent-antibody (IFA) assay with antigen produced in vitro in the human promyelocytic leukemia cell line HL60 was used to identify the first recognized case of human granulocytic ehrlichiosis in Rhode Island. This IFA assay was used to detect granulocytic ehrlichiae in white-footed mice and in a dog inhabiting the area surrounding the patient's residence. Host-seeking Ixodes scapularis ticks found in the same habitat also were infected. I. scapularis ticks collected from other locations were fed on dogs and New Zealand White rabbits to assess the competency of these species as hosts of granulocytotropic Ehrlichia. Tick-induced infections of dogs were confirmed by serologic testing, tissue culture isolation, and PCR amplification, whereas several rabbits seroconverted but were PCR and culture negative. PCR amplification of the 16S rRNA gene and DNA sequencing of the PCR products or culture isolation was used to confirm granulocytic Ehrlichia infections in humans, dogs, white-footed mice, and ticks.
Molecular characterization of the gene encoding H antigen in Escherichia coli and development of a PCR-restriction fragment length polymorphism test for identification of E. coli O157:H7 and O157:NM
Authors: Fields P I , Blom K , Hughes H J , Helsel L O , Feng P , Swaminathan B
Published: January 2020

Recent outbreaks of disease caused by Escherichia coli O157:H7 have focused much attention on this newly emerged pathogen. Identification of the H7 flagellar antigen is critical for the confirmation of E. coli O157:H7; however, clinical isolates are frequently nonmotile and do not produce detectable H antigen. To further characterize nonmotile isolates (designated NM), we developed a PCR-restriction fragment length polymorphism (PCR-RFLP) test to identify and characterize the gene encoding the H antigen (fliC) in E. coli. The entire coding sequence of fliC was amplified by PCR, the amplicon was restricted with RsaI, and the restriction fragment pattern was examined after gel electrophoresis. Two hundred eighty E. coli isolates representing serotypes O157:H7 and O157:NM, flagellar antigen H7 groups associated with other O serogroups, and all other flagellar antigen groups were analyzed. A single restriction pattern (pattern A) was identified for O157:H7 isolates, O157:NM isolates that produced Shiga toxin (formerly Shiga-like toxin or verotoxin), and 16 of 18 O55:H7 isolates. Flagellar antigen group H7 isolates of non-O157 serotypes had one of three banding patterns distinct from pattern A. A wide variety of patterns were found among isolates of the other 52 flagellar antigen groups; however, none was identical to the O157:H7 pattern. Thirteen of 15 nonmotile strains that did not produce the A pattern had patterns that matched those of other known H groups. The PCR-RFLP in conjunction with O serogroup determination will be useful in identifying E. coli O157:H7 and related strains that do not express immunoreactive H antigen and could be expanded to include other clinically important E. coli strains.