A model to determine at-home restrictions for cats after treatment of hyperthyroidism with radioiodine.

OBJECTIVES: Felinehyperthyroidism is the most common endocrine disease of older cats and radioiodine is considered to be the gold standard treatment. Isolation periods following treatment vary depending on both individual treatment facilities and the relevant legislation of the country; therefore, there is no recognised standardised protocol defining the length of isolation. This work describes how our institution validated that its owner restrictions met dose constraints by using a model of iodine retention to calculate the required duration and nature of owner restrictions. MATERIALS AND METHODS: The retained radioactivity of cats at the point of discharge was used to simulate the radiation dose to owners in the 90 days following release. The model created was used to calculate the minimum duration of isolation for a range of administered activities and owner restrictions. RESULTS: Using the model, it was found that when injected with the maximum dose used, 222 MBq radioiodine, it was possible to release cats after 14 days of isolation and keep owner doses below 0.30 mSv (whole-body effective dose constraint for a single radiation source) with some restrictions. It was possible to release after 23 days with no restrictions. CLINICAL SIGNIFICANCE: The present study provides clinicians with a consistent and verified method in which they can calculate the isolation periods for radioiodine-treated cats.

Dorsal mediastinal haemorrhage in dogs: 4 cases (2010-2018).

OBJECTIVE: This retrospective case series describes the clinical presentation and CT findings of dogs with presumed mediastinal haemorrhage with no apparent identifiable underlying cause. MATERIALS AND METHODS: Medical records were searched for dogs with presumed or suspected mediastinal haemorrhage of non-thymic origin. For all dogs, data on signalment, history, physical examination, treatment and outcome were collected by reviewing the medical records. Follow-up information was collected by telephone interviews with the owners and/or their primary-care veterinarians. RESULTS: Four dogs were included. All survived to discharge with apparent resolution of the mediastinal haemorrhage (based on repeat imaging and/or clinical signs) with supportive treatment alone. Follow-up information was available from 2 months to 5 years following discharge, and none of the patients showed a recurrence of clinical signs during this period. CLINICAL SIGNIFICANCE: This case series highlights that presumed haemorrhage into the mediastinum can occur in dogs without an obviously identifiable cause and, whilst rare, should be considered as a cause of dorsal mediastinal masses and may be successfully managed with supportive care alone.

Molecular typing and resistance analysis of travel-associated Salmonella enterica serotype Typhi.

Salmonella enterica serotype Typhi is a human pathogen causing 12 to 30% mortality and requiring antibiotic therapy to control the severity of the infection. Typhoid fever in United States is often associated with foreign travel to areas of endemicity. Increasing resistance to multiple drugs, including quinolones, is associated with decreased susceptibility to ciprofloxacin (DCS). We investigated 31 clinical strains isolated in Florida from 2007 to 2010, associated with travel to six countries, to examine the clonal distribution of the organism and apparent nalidixic acid (NAL) resistance. The strains were isolated from blood or stool of patients aged 2 to 68 years. The isolates were subtyped by ribotyping and pulsed-field gel electrophoresis. Susceptibilities to 15 antimicrobials were determined, and the isolates were screened for integrons and gyrase A gene mutations. Both typing techniques effectively segregated the strains. Identical clones were associated with different countries, while diverse types coexisted in the same geographic location. Fifty-one percent of the strains were resistant to at least one antimicrobial, and five were resistant to three or more drugs (multidrug resistant [MDR]). All 12 isolates from the Indian subcontinent were resistant to at least one drug, and 83% of those were resistant to NAL. Three of the MDR strains harbored a 750-bp integron containing the dfr7 gene. Ninety-three percent of the resistant strains showed a DCS profile. All the NAL-resistant strains contained point mutations in the quinolone resistance-determining region of gyrA. This study affirms the global clonal distribution, concomitant genetic heterogeneity, and increased NAL resistance of S. enterica serovar Typhi.

Bacillus Strains Most Closely Related to Bacillus nealsonii Are Not Effectively Circumscribed within the Taxonomic Species Definition.

Bacillus strains with >99.7% 16S rRNA gene sequence similarity were characterized with DNA:DNA hybridization, cellular fatty acid (CFA) analysis, and testing of 100 phenotypic traits. When paired with the most closely related type strain, percent DNA:DNA similarities (% S) for six Bacillus strains were all far below the recommended 70% threshold value for species circumscription with Bacillus nealsonii. An apparent genomic group of four Bacillus strain pairings with 94%-70% S was contradicted by the failure of the strains to cluster in CFA- and phenotype-based dendrograms as well as by their differentiation with 9-13 species level discriminators such as nitrate reduction, temperature range, and acid production from carbohydrates. The novel Bacillus strains were monophyletic and very closely related based on 16S rRNA gene sequence. Coherent genomic groups were not however supported by similarly organized phenotypic clusters. Therefore, the strains were not effectively circumscribed within the taxonomic species definition.

Survival and the development of azotemia after treatment of hyperthyroid cats.

BACKGROUND: Hyperthyroidism complicates the diagnosis of chronic kidney disease (CKD) as it increases glomerular filtration rate. No practical and reliable means for identifying those cats that will develop azotemia after treatment for hyperthyroidism has been identified. Hyperthyroidism is associated with proteinuria. Proteinuria has been correlated with decreased survival of cats with CKD and with progression of CKD. HYPOTHESIS: Proteinuria and other clinical parameters measured at diagnosis of hyperthyroidism will be associated with the development of azotemia and survival time. ANIMALS: Three hundred client owned hyperthyroid cats treated in first opinion practice. METHODS: Retrospective, cohort study relating clinical parameters in hyperthyroid cats at diagnosis to the development of azotemia within 240 days of diagnosis and survival time (all cause mortality). Multivariable logistic regression analysis was used to identify factors that were predictive of the development of azotemia. Multivariable Cox regression analysis was used to identify factors associated with survival. RESULTS: Three hundred cats were eligible for survival analysis and 216 cats for analysis of factors associated with the development of azotemia. The median survival time was 417 days, and 15.3% (41/268) cats developed azotemia within 240 days of diagnosis of hyperthyroidism. Plasma concentrations of urea and creatinine were positively correlated with the development of azotemia. Plasma globulin concentration was negatively correlated with the development of azotemia. Age, urine protein:creatinine ratio, and the presence of hypertension were significantly correlated with decreased survival time. Urine specific gravity and PCV were significantly correlated with increased survival time. CONCLUSIONS AND CLINICAL IMPORTANCE: The proteinuria associated with hyperthyroidism is not a mediator of progression of CKD; however, it does correlate with all cause mortality.

Two-year study evaluating the potential loss of methicillin resistance in a methicillin-resistant Staphylococcus aureus culture collection.

A reported loss of mecA prompted us to monitor 360 cryostocked methicillin-resistant Staphylococcus aureus strains for stability. Concurrently, 14 well-characterized strains were stored in a Microbank preservation system and subjected to multiple freeze-thaw events. There were no significant declines in the methicillin-resistant populations with either method over a two-year period.

Fatal necrotizing pneumonia due to a Panton-Valentine leukocidin positive community-associated methicillin-sensitive Staphylococcus aureus and Influenza co-infection: a case report.

Recent studies have described a number of fatalities due to methicillin-resistant Staphylococcus aureus (MRSA) and influenza virus co-infection. MRSA isolates provide a challenge to caregivers due to inherent wide range antibiotic resistance. Many facilities have instituted screening methods, based on the presence of antibiotic resistance genes, to identify MRSA positive patients upon admission. However, the resistance profile of the pathogen does not necessarily determine the severity of disease caused by that organism. We describe a fatal case of necrotizing pneumonia in a patient co-infected with Influenza B and a community-associated, PVL-positive methicillin-susceptible Staphylococcus aureus (MSSA).

Molecular characterization of Vibrio parahaemolyticus strains associated with foodborne illness in Florida.

Vibrio parahaemolyticus is the leading cause of bacterial seafood-based illness in the United States. Real-time PCR, pandemic group-specific PCR, ribotyping, and multilocus sequence typing were used to characterize 30 strains of V. parahaemolyticus including 11 strains associated with foodborne outbreaks in Florida and 6 known pandemic strains. Thirteen strains were positive for four pandemic group-specific PCR markers, including 5 strains associated with outbreaks in Florida. Molecular typing methods were used to further define the pandemic status of these strains because the current PCR markers are not sufficient to identify pandemic isolates. Nine of the Florida strains clustered with a majority of the known pandemic strains, based on ribotyping patterns using PvuII, but no isolated pandemic branch was formed. Using multilocus sequence typing, it was determined that 14 strains possess a previously determined pandemic sequence type. This study identified 13 novel sequence types and seven to nine novel alleles for each locus. Furthermore, the results indicate that seven of the strains from recent foodborne outbreaks in Florida are pandemic strains, and that multilocus sequence typing was the most accurate molecular method to identify these strains.

Bacillus acidiceler sp. nov., isolated from a forensic specimen, containing Bacillus anthracis pX02 genes.

Research at the Center for Biological Defense identified plasmid-borne forms of Bacillus anthracis pXO2 genes in a Gram-positive, endospore-forming rod, isolated from a forensic specimen considered a credible threat of harbouring anthrax. Conventional, commercial and molecular-based methods indicated that the isolate (CBD 119(T)) was not B. anthracis and considered not to be a member of the Bacillus cereus group. Based on the 16S rRNA gene sequence similarities, strain CBD 119(T) was most closely related to Bacillus luciferensis LMG 18422(T) (99.3 %). Phenotyping and fatty acid methyl ester analysis of the isolate were conducted alongside B. luciferensis JCM 12212(T). The major cellular fatty acids (anteiso-C(15 : 0), iso-C(15 : 0), and >7 iso or anteiso forms) supported inclusion of the isolate in the genus Bacillus. Strain CBD 119(T) was inconsistent with B. luciferensis JCM 12212(T) for 18 of 96 traits evaluated including motility, degree of endospore-driven swelling and pH optimum; the two were linked by fatty acid methyl ester analysis as separate but closely related species. DNA-DNA relatedness between strain CBD 119(T) and B. luciferensis JCM 12212(T) resulted in less than 20 % hybridization. The results of biochemical and physiological characterization, chemotaxonomic analysis and DNA-DNA hybridization differentiated strain CBD 119(T) both phenotypically and genotypically from the only species with validly published name with greater than 97 % 16S rRNA gene sequence similarity. The isolate has an accelerated doubling time when grown in aerated broth at pH 5.9 relative to that at pH 7.1. Therefore, it is proposed that strain CBD 119(T) represents a novel species, Bacillus acidiceler sp. nov. The type strain is strain CBD 119(T) (=NRRL B-41736(T)=DSM 18954(T)).

Bacillus anthracis virulent plasmid pX02 genes found in large plasmids of two other Bacillus species.

In order to cause the disease anthrax, Bacillus anthracis requires two plasmids, pX01 and pX02, which carry toxin and capsule genes, respectively, that are used as genetic targets in the laboratory detection of the bacterium. Clinical, forensic, and environmental samples that test positive by PCR protocols established by the Centers for Disease Control and Prevention for B. anthracis are considered to be potentially B. anthracis until confirmed by culture and a secondary battery of tests. We report the presence of 10 genes (acpA, capA, capB, capC, capR, capD, IS1627, ORF 48, ORF 61, and repA) and the sequence for the capsule promoter normally found on pX02 in Bacillus circulans and a Bacillus species closely related to Bacillus luciferensis. Tests revealed these sequences to be present on a large plasmid in each isolate. The 11 sequences consistently matched to B. anthracis plasmid pX02, GenBank accession numbers AF188935.1, AE011191.1, and AE017335.3. The percent nucleotide identities for capD and the capsule promoter were 99.9% and 99.7%, respectively, and for the remaining nine genes, the nucleotide identity was 100% for both isolates. The presence of these genes, which are usually associated with the pX02 plasmid, in two soil Bacillus species unrelated to B. anthracis alerts us to the necessity of identifying additional sequences that will signal the presence of B. anthracis in clinical, forensic, and environmental samples.

Community-associated methicillin-resistant Staphylococcus aureus epidemic clone USA300 in isolates from Florida and Washington.

We examined 299 methicillin-resistant, community-associated Staphylococcus aureus isolates from Florida and Washington State for the presence of the USA300 epidemic clone. Pulsed-field gel electrophoresis demonstrated the epidemic clone in 43% of our S. aureus strains and in isolates from both states. The majority of the USA300 isolates (88%) were from wound infections.

Use of two selective media and a broth motility test can aid in identification or exclusion of Bacillus anthracis.

During the anthrax attack of 2001, the Florida Department of Health (FDOH) Bureau of Laboratories in Tampa received hundreds of isolates suspected of being Bacillus anthracis. None were confirmed to be B. anthracis since most isolates were motile and not even in the Bacillus cereus group. Although the sentinel laboratories now send fewer isolates to FDOH laboratories, should another attack occur the number of isolates submitted would likely increase dramatically, and this upsurge would seriously challenge personnel who are expected to be busy examining an increased number of environmental samples. We examined two selective and differential growth media and alternative motility methods that could be used to streamline the processing of suspicious isolates. Of 60 isolates previously sent to the FDOH laboratory, 56 were endospore-forming gram-positive rods and only 7 grew on mannitol-egg yolk-polymyxin B agar and/or the Anthracis chromogenic agar. Microscopic observation of early-log-phase growth (2 to 3 h) in a shaking broth was the best method to detect motility in 40 isolates that appeared nonmotile in the motility media investigated. One of these growth media and microscopic examination of shaken broth cultures can be used to show that an isolate is not B. anthracis before expensive molecular and antibody-based tests are performed. By doing so, costs could be reduced and analysis time shortened.

Real-time PCR detection of the thermostable direct hemolysin and thermolabile hemolysin genes in a Vibrio parahaemolyticus cultured from mussels and mussel homogenate associated with a foodborne outbreak.

Molecular methods have become vital epidemiological tools in the detection and characterization of bacteria associated with a foodborne outbreak. We used both culture and real-time PCR to detect a Vibrio parahaemolyticus isolate associated with a foodborne outbreak. The outbreak occurred in July 2002 in Polk County, Florida, and originated at a Chinese buffet, with one person being hospitalized. The hospital isolated V. parahaemolyticus from the patient but destroyed the sample after diagnosis. From an onsite visit of the restaurant, food samples that possibly contributed to the outbreak were collected and sent to the Florida Department of Health, Tampa Branch Laboratory. Crab legs, crabsticks, and mussel samples were homogenized and incubated according to the Food and Drug Administration Bacteriological Analytical Manual culture protocol. Three sets of primers and a TaqMan probe were designed to target the tdh, trh, and tlh genes and used for real-time PCR. This study was successful in isolating V. parahaemolyticus from a mussel sample and detecting two of its genes (tdh and tlh) in food and pure culture by real-time PCR.

Comparison of methods for DNA isolation from food samples for detection of Shiga toxin-producing Escherichia coli by real-time PCR.

In this study, food samples were intentionally contaminated with Escherichia coli O157:H7, and then DNA was isolated by using four commercial kits. The isolated DNA samples were compared by using real-time PCR detection of the Shiga toxin genes. The four kits tested worked similarly.

A rapid and efficient assay for extracting DNA from fungi.

AIMS: A method for the rapid extraction of fungal DNA from small quantities of tissue in a batch-processing format was investigated. METHODS AND RESULTS: Tissue (< 3.0 mg) was scraped from freshly-grown fungal isolates. The tissue was suspended in buffer AP1 and subjected to seven rounds of freeze/thaw using a crushed dry ice/ethanol bath and a boiling water bath. After a 30 min boiling step, the tissue was quickly ground against the wall of the microfuge tube using a sterile pipette tip. The Qiagen DNeasy Plant Tissue Kit protocol was then used to purify the DNA for PCR/sequencing applications. CONCLUSIONS: The method allowed batch DNA extraction from multiple fungal isolates using a simple yet rapid and reliable assay. SIGNIFICANCE AND IMPACT OF THE STUDY: Use of this assay will allow researchers to obtain DNA from fungi quickly for use in molecular assays that previously required specialized instrumentation, was time-consuming or was not conducive to batch processing.

Design and synthesis of alpha,beta-unsaturated carbonyl compounds as potential ACE inhibitors.

The alpha,beta-unsaturated amide that is incorporated into the basic structural frame of a simple substrate molecule of angiotensin converting enzyme was found to serve as a Michael acceptor for the catalytic carboxylate of Glu-127, inhibiting the enzyme irreversibly.