Identification of Brucella abortus biovar 4 of bovine origin in Colombia / Identificación de Brucella abortus biovar 4 de origen bovino en Colombia

The objective of this study was to identify twelve Brucella abortus isolates of bovine origin from the department of Nariño in Colombia up to the biovar level. These isolates are included in the collection of the Germplasm Bank of Microorganisms of Animal Health Interest -Bacteria and Virus (BGSA-BV). The identification was carried out through conventional methods such as macro and microscopic morphological descriptions, enzymatic activity, biochemical profile, substrate use and sensitivity to dyes. Complementary genotypic characterization was carried out using multiplex PCR for B. abortus, Brucella melitensis, Brucella ovis, and Brucella suis-Erytritol (AMOS-ERY-PCR), RFLP-IS711, by southern blot hybridization, as well as by the multiple locus variable number of tandem repeat analysis (MLVA) using the ery gene and the insertion sequence IS711 and variable number of tandem repeats (VNTR) as molecular markers. The results of the phenotypic and molecular characterization allowed to identify twelve isolates as B. abortus biovar 4 as well as to differentiate field from vaccine strains. This is the first study on the phenotypic and molecular identification of B. abortus isolates in Colombia. It was concluded that the phenotypic and molecular identification of twelve isolates as B. abortus biovar 4 could be achieved using conventional and molecular techniques with enough resolution power. The identification of these isolates to the biovar level in taxonomic and epidemiological terms will allow the use of this genetic resource as reference strains in future research. This finding constitutes the basis for identifying biotypes not previously reported in the country that might be useful to support brucellosis survey programs in Colombia.

El objetivo de este estudio fue identificar 12 aislamientos de Brucella abortus de origen bovino procedentes del departamento de Narino, Colombia, hasta la descripción de biovar. Estos aislamientos conforman la colección del Banco de Germoplasma de Microorganismos de Interés en Salud Animal, Bacterias y Virus. La identificación se hizo mediante métodos convencionales, como la descripción morfológica macro y microscópica de actividad enzimática, de perfiles bioquímicos, de utilización de sustratos y de sensibilidad a colorantes. Se hizo una caracterización genotipica complementaria mediante PCR múltiple para Brucella abortus, Brucella melitensis, Brucella ovisy Brucella suis-eritritol (AMOS-ERY-PCR); RFLP-/S7II; hibridación Southern blot y análisis multi-locus de repeticiones en tándem de número variable (MLVA), empleando como marcadores moleculares el gen ery, la secuencia de inserción /S711 y el número variable de repeticiones en tándem (VNTR). Los resultados de la caracterización fenotípica y molecular permitieron identificar 12 aislamientos de campo como B. abortus biovar 4 y diferenciar cepas de campo de cepas vacunales. Este es el primer estudio de identificación fenotípica y molecular de aislamientos de B. abortus en Colombia. Por su importancia taxonómica y epidemiológica, la identificación de estos aislamientos hasta el nivel de biovar permitirá disponer de recursos genéticos que se pueden emplear como cepas de referencia en futuras investigaciones. Estos resultados pueden considerarse como una base para la identificación de biotipos no reportados en el país y podrán ser utilizados en programas de monitoreo y vigilancia de la brucelosis bovina en Colombia.

A unique DNA repair and recombination gene (recN) sequence for identification and intraspecific molecular typing of bacterial wilt pathogen Ralstonia solanacearum and its comparative analysis with ribosomal DNA sequences.

Ribosomal gene sequences are a popular choice for identification of bacterial species and, often, for making phylogenetic interpretations. Although very popular, the sequences of 16S rDNA and 16-23S intergenic sequences often fail to differentiate closely related species of bacteria. The availability of complete genome sequences of bacteria, in the recent years, has accelerated the search for new genome targets for phylogenetic interpretations. The recently published full genome data of nine strains of R. solanacearum, which causes bacterial wilt of crop plants, has provided enormous genomic choices for phylogenetic analysis in this globally important plant pathogen. We have compared a gene candidate recN, which codes for DNA repair and recombination function, with 16S rDNA/16-23S intergenic ribosomal gene sequences for identification and intraspecific phylogenetic interpretations in R. solanacearum. recN gene sequence analysis of R. solanacearum revealed subgroups within phylotypes (or newly proposed species within plant pathogenic genus, Ralstonia), indicating its usefulness for intraspecific genotyping. The taxonomic discriminatory power of recN gene sequence was found to be superior to ribosomal DNA sequences. In all, the recN-sequence-based phylogenetic tree generated with the Bayesian model depicted 21 haplotypes against 15 and 13 haplotypes obtained with 16S rDNA and 16-23S rDNA intergenic sequences, respectively. Besides this, we have observed high percentage of polymorphic sites (S 23.04%), high rate of mutations (Eta 276) and high codon bias index (CBI 0.60), which makes the recN an ideal gene candidate for intraspecific molecular typing of this important plant pathogen.

Investigation of Ureaplasma urealyticum biovars and their relationship with antimicrobial resistance.

Purpose: To develop Taqman fluorescence quantitative polymerase chain reaction (PCR) method for investigating the characteristics of the distributions of Ureaplasma urealyticum (UU) biovars and to explore the relationship between UU biovars and antimicrobial resistance. Materials and Methods: By the method of culture, Ureaplasma species were detected. Taqman fluorescence quantitative PCR for detecting UU biovars were developed and UU clinical isolates were detected to distinguish biovars. The broth micro-dilution susceptibility testing methods were used to determine UU susceptibility. Results: By Taqman PCR method, UU biovars was successfully detected. Of 126 samples, biovar 1 was found in 73 (57.94%). There was a statistical difference between genital-urinary tract infection group and asymptomatic group (P<0.05). In the region, UU biovar 1 to 9 kinds of agents kept higher susceptibility rates, but biovar 2 maintained higher susceptibility rates only to tetracyclines. Compared with biovar 1, UU biovar 2 resistance rates to 7 kinds of agents were higher (P<0.05). Conclusions: (1) Our new established Taqman PCR method is a useful tool for screening UU biovars. (2) UU biovar 1 predominated in asymptomatic population; whereas in genital-urinary tract infection population UU biovar 2 had a higher proportion. (3) The characteristics of drug resistance were different between UU biovars. Overall, both two biovars remained higher susceptibility rates to tetracyclines. A majority of biovor 1 strains were sensitive to macrolides and quinolones; while only a small number of biovar 2 strains kept sensitive to roxithromycin and quinolones, a large proportion of biovar 2 strains were found in intermediate ranges.

Different Clinical Characteristics Among Aeromonas hydrophila, Aeromonas veronii biovar sobria and Aeromonas caviae Monomicrobial Bacteremia

This study aimed to compare the clinical presentations of Aeromonas hydrophila, A. veronii biovar sobria and A. caviae monomicrobial bacteremia by a retrospective method at three hospitals in Taiwan during an 8-yr period. There were 87 patients with A. hydrophila bacteremia, 45 with A. veronii biovar sobria bacteremia and 22 with A. caviae bacteremia. Compared with A. hydrophila and A. veronii biovar sobria bacteremia, A. caviae bacteremia was more healthcare-associated (45 vs 30 and 16%; P = 0.031). The patients with A. caviae bacteremias were less likely to have liver cirrhosis (27 vs 62 and 64%; P = 0.007) and severe complications such as shock (9 vs 40 and 47%; P = 0.009) and thrombocytopenia (45 vs 67 and 87%; P = 0.002). The APACHE II score was the most important risk factor of Aeromonas bacteremia-associated mortalities. The APACHE II scores of A. caviae bacteremias were lower than A. hydrophila bacteremia and A. veronii biovar sobria bacteremia (7 vs 14 and 16 points; P = 0.002). In conclusion, the clinical presentation of A. caviae bacteremia was much different from A. hydrophila and A. veronii biovar sobria bacteremia. The severity and mortality of A. caviae bacteremia were lower than A. hydrophila or A. veronii biovar sobria bacteremia.

Molecular Epidemiology of Ureaplasma urealyticum in Patients with Urogenital Infections / 中华医院感染学杂志

OBJECTIVE To investigate the Ureaplasma urealyticum infection in patients with urogenital infections and the distribution of its serotypes and genotypes.METHODS The clinical samples were firstly screened with B-Merieux mycoplasma ID2 culture and identification kit,then the positive cultural samples were subtyped by PCR method.RESULTS The positive rate of U.urealyticum with cultural kit was 37.43%(670/1790).The 392 positive cultural samples were subtyped with PCR,the positive rate of U.parvum was 71.94% and the positive rate of U.urealyticum was 29.85%.The serotypes of 264 U.parvum positive samples were identified with PCR,there were 20 samples with serovar 1(7.58%),139 samples with serovar 3 or 14(52.65%),and 51 samples with serovar 6(19.32%).Ninety nine positive samples with U.urealyticum were genotyped,the results showed that the rate of genotype 3(serovar 7 or 11) was lower than genotypes 1 and 2.CONCLUSIONS PCR identification and subtyping of Ureaplasma in patients with urogenital infections showed that the major epidemic pathogen of Ureaplasma in east of Guangdong Province is U.urealyticum serovar 3 and serovar 6,and the distribution of different serotypes or genotypes of Ureaplasma in this area is specific.

Misidentification as Vibrio alginolyticus with Vitek GNI+ Card in Three Cases / 대한임상미생물학회지

The introduction of a new, fully automated system into the clinical microbiology laboratory contributes to a rapid identification of microorganisms with accurate and reliable results, but such a system requires a high cost and additional tests for identification of some species. For instance, additional tests on oxidase, indole, motility, hemolysis, and pigmentation are needed in the correct identification by using Vitek GNI+ system (bioMerieux Vitek Inc., MO, USA). In particular, Vibrio and Aeromonas species are occasionally identified incorrectly when an automated system is used, and thus conventional biochemical tests may be more reliable in the identification of such species. We experienced three cases of incorrect identification of Vibrio parahaemolyticus, Vibrio cholerae, and Aeromonas veronii biovar sobria as Vibrio alginolyticus by using Vitek GNI+ card.

Molecular mechanism of biovar conversion of Yersinia pestis and the proposition of a new biovar:Microtus / 解放军医学杂志

Objective To study the genetic basis for biovar conversion of Y. pestis. Methods In silico comparative genomic analysis was conducted and some critical genetic variations of Yersinia pestis were comparatively analyzed by means of PCR and DNA sequencing. Results A 93bp in-frame deletion in glpD gene results in the glycerol negative characteristic of Orientalis strains. A point mutation in the napA gene may cause the negative characteristic of nitrate reduction in Mediaevalis and Microus strains. A 122-bp frameshift deletion in the araC gene may lead to the arabinose negative phenotype of Microus strains. Conclusion In this study, Microtus strains with their unique pathogenic, biochemical and molecular features, were proposed as a novel biovar Microtus. In the light of its differential ability to ferment glycerol and arabinose and to reduce nitrate, Y. pestis can be classified into four biovars-Antiqua(glycerol positive, arabinose positive and nitrate positive), Mediaevalis(glycerol positive, arabinose positive and nitrate negative), Orientalis(glycerol negative, arabinose positive and nitrate positive), and Microtus(glycerol positive, arabinose negative and nitrate negative).