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PubMed1992

Detection and serotyping of rotaviruses in stool specimens by using reverse transcription and polymerase chain reaction amplification.

Direct rotavirus serotyping (VP7, G type) in stool specimens was carried out by reverse transcription and polymerase chain reaction amplification (RT-PCR) and compared to serotyping by enzyme immunoassay with monoclonal antibodies (EIA-MAb). The methods used for double-stranded (ds) RNA extraction, RT-PCR amplification, and the primers used were modified from previous reports [Gouvea et al.: Journal of Clinical Microbiology 29:519-523, 1990; Gentsch et al.: Journal of Clinical Microbiology, 1992]. For samples that were positive by both methods, the serotypes obtained were identical, however RT-PCR typing was found to be considerably more sensitive (70.4% samples serotyped) than EIA-MAb (35.6% of samples serotyped). The overall sensitivities for detection of rotavirus in stool samples by latex agglutination, enzyme immunoassay, electron microscopy, polyacrylamide gel electrophoresis, and RT-PCR were essentially the same. The results confirm that RT-PCR typing (genotyping) is extremely valuable for G typing of samples which cannot be typed by EIA-MAb. We also developed a PCR confirmation technique for serotypes 1, 2, and 4.

Acute Disease↗
PubMed1999

A multiplex PCR for Massachusetts and Arkansas serotypes of infectious bronchitis virus.

Infectious bronchitis virus (IBV), the prototype of the coronavirus family, is an enveloped, single-stranded RNA virus with a genome size of approximately 27.6 kilobase. Infectious bronchitis virus causes an acute, highly contagious respiratory and urogenital disease of chickens which results in significant economic losses in commercial broilers, layers and breeders. A rapid, highly sensitive and specific method is needed in the differential diagnosis of infections of different serotypes. A multiplex polymerase chain reaction (PCR) method was developed and optimized to simultaneously detect Massachusetts (Mass) and Arkansas (Ark) serotypes of IBV. One common primer and two serotype specific primers were chosen from the S1 gene sequences of IBV and used in one PCR reaction. Under optimized PCR conditions, two serotype specific PCR products, 1026 bp for Mass and 896 bp for Ark, respectively, were amplified and detected by agarose gel electrophoreses. The specificity of the technique was verified by using 20 different strains and isolates of IBV, and other avian bacterial and viral pathogens. Using a serial 10-fold dilution of the artificial mixture of both Mass and Ark samples, the detection limit was found to be 5 pg RNA after 35 cycles of PCR. The multiplex PCR was able to detect and differentiate both serotypes in embryonated eggs that were co-infected with different EID50virus titers of Mass 41 and Ark 99. The multiplex PCR developed in this study will be valuable for rapid identification, differential diagnosis, and epidemiological studies of these two serotypes of IBV infections.

Animals↗
PubMed1993

DNA sequence analysis of an allelic variant of the Actinobacillus pleuropneumoniae-RTX-toxin I (ApxIA) from serotype 10.

The structural gene encoding Actinobacillus pleuropneumoniae-RTX-toxin I (ApxIA), one of the major hemolytic and cytolytic toxins of the organism, was cloned from serotype 10. The nucleotide sequence data showed that the gene from serotype 10 was 98% identical to that from serotype 1 at both DNA and amino acid levels. The sequence difference was found to localize at the 3' terminal region of the gene. We then analyzed the 3' terminal region of the apxIA gene from other serotypes, 5a, 5b, 9 and 11, using polymerase chain reaction-amplified DNA fragments. Results of DNA sequence indicated that apxIA gene can be divided into the original form including serotypes 1, 9 and 11, and the allelic variant including serotypes 5a, 5b and 10. These gene products, ApxIA proteins, appear to have different second structures at the carboxyl terminal proximal region.

Actinobacillus pleuropneumoniae↗
PubMed1994

Identification of a new VP4 serotype of human rotaviruses.

A new VP4 gene allele was identified by sequencing of two rotavirus recovered from infants with diarrhea. Strains HAL1166 (a VP7 serotype 8 human rotavirus from Finland) and PA169 (a VP7 serotype 6 human rotavirus from Italy) present 22-43% nucleotide sequence divergence and 14-45% amino acid divergence in the VP4 gene with rotaviruses belonging to various VP4 gene alleles previously reported. Cross-neutralization analysis between PA169 and rotaviruses belonging to the other recognized VP4 (P) serotypes had previously shown that none of them was similar to PA169. Hyperimmune serum prepared against a reassortant with the VP4 gene of strain PA169 and the VP7 gene of serotype 2 strain DS1 failed to neutralize previously recognized human rotavirus VP4 serotypes, indicating that PA169 represents a new VP4 serotype specificity. In addition, one of four neutralizing monoclonal antibodies directed to the VP4 of PA169 was specific for this virus, while three others had very limited cross-reactivity with rotaviruses possessing different VP4s.

Amino Acid Sequence↗
PubMed1996

Classification of rotavirus VP4 and VP7 serotypes.

Rotaviruses, members of the Reoviridae family, are major etiologic agents of acute nonbacterial gastroenteritis of the young in a wide variety of mammalian and avian species, including humans. The need for effective immunoprophylaxis against rotaviral gastroenteritis has stimulated interest in the biochemical, molecular, genetic, and clinical aspects of these agents with the aim of developing safe and effective vaccines. Because neutralizing antibodies appear to play an important role in protection against many viral diseases, rotavirus antigens that induce neutralizing antibodies have played a central role in research and development of a rotavirus vaccine. The VP7 glycoprotein and VP4 spike protein that constitute the outer capsid of a complete rotavirus particle have been shown to be independent neutralization antigens. Since type specificity of the outer capsid proteins of a rotavirus appears to play an important role in protection against disease in experimental animal models, continued efforts have been made for classification and typing of neutralization specificities on the VP7 or VP4 capsid protein. Based on a criterion of > 20-fold differences between the homologous and heterologous reciprocal neutralizing antibody titers, fourteen VP7 (G) serotypes have been established. Studies are underway to characterize and classify the VP4 (P) serotypes among the strains that exhibit the fourteen different G serotypes. Attempts to classify the VP4 serotypes based on the same criterion (i.e., > 20-fold antibody differences) that is applied to classification of VP7 serotypes are in progress. This standard of > 20-fold antibody differences can be applied with hyperimmune serum raised to a reassortant possessing the VP4 encoding gene (and an unrelated VP7 encoding gene). Genotypes can provide leads towards classification but the serotype of a strain should be based on neutralization.

Animals↗
PubMed1993

The isolation of Cryptococcus neoformans from pigeon droppings and serotyping of naturally and clinically sourced isolates in China.

This is the first report on the isolation of Cryptococcus neoformans from pigeon droppings in China and their serotypes. C. neoformans colonies which produced brown colonies on caffeic acid-cornmeal agar were found in Twenty-five out of thirty-six samples of pigeon droppings. Fifty-one colonies randomly picked from the positive samples were identified as C. neoformans by a commercially available kit for carbon source assimilation test and Christensen's urea agar. Forty (78%) out of the 51 strains were serotyped as A and 11 (22%) as AD. At the same time, seventeen out of nineteen clinical isolates were serotyped as A and 2 as B. There are three findings in our results. One is that only C. neoformans var. neoformans strains could be isolated from pigeon droppings, although the variety gattii strains were found in the clinical isolates obtained in the same geographic site in China. The second is that serotype A strains were most frequently seen in natural and clinical materials in the southeast part of China, and serotype AD strains were isolated in pigeon droppings but not in clinical materials. The third is that the coexistence of serotype A and AD cells of C. neoformans strains in same samples of pigeon droppings were observed.

Animals↗
PubMed1985

Detection and prevalence of serotypes of feline syncytial spumaviruses.

Three serotypes of feline syncytial virus (FSV) were detected by neutralisation tests: 906, a serotype of low prevalence and 702 and 951 which were serotypes of higher prevalence, between which a minor one-way antigenic difference was detected. Serum antibody in naturally-infected cats in some cases neutralised 951 but not 702 or 906 which suggested that 951 could be considered as a major distinct serotype. An increase in prevalence of antibody to FSV in cats over a 5 year period from 1977-1981 was detected by neutralisation, agar gel immunodiffusion, and fluorescent antibody techniques. Over the 5 year period the prevalence of antibody to the 951 serotype increased and the overall increase in prevalence of antibody to FSV during this period appeared to relate to dissemination of the 951 serotype.

Animals↗
PubMed1989

Epidemiology of rotavirus strains infecting children throughout Australia during 1986-1987. A study of serotype and RNA electropherotype.

The epidemiology of human rotaviruses throughout Australia was studied by examining 344 rotavirus positive faecal specimens using an enzyme immunoassay incorporating serotype specific monoclonal antibodies. Specimens were collected from children less than 5 years old admitted to urban hospitals for treatment of acute diarrhoea during the winter months of 1986 and/or 1987 in Queensland, New South Wales, Victoria, Tasmania, South Australia, and Western Australia. The infecting rotavirus serotype was identified in 229 of 344 (66.6%) specimens. The predominant serotype throughout Australia was serotype 1 which was identified in 218 of 229 (95%) typable specimens. The majority (201 of 218) were identified as monotype 1a strains. Serotype 2 strains were found in Perth, Western Australia in 2 of 12 specimens collected in 1986 and in 6 of 32 specimens collected in 1987. RNA electropherotypes comprising 30 different patterns were detected after co-electrophoresis of 143 of the 218 serotyped strains. Twenty-nine electropherotypes were serologically homogeneous. One electropherotype contained strains with different monotypes including 1a, 1b, and 1d. The results show remarkable serological uniformity, associated with genetic diversity of rotavirus strains identified in widely separated areas of Australia during one winter.

Australia↗
PubMed1990

Isolation in Europe of 69 M-like (serotype 8) human rotavirus strains with either subgroup I or II specificity and a long RNA electropherotype.

During an epidemiological study on the prevalence of human rotavirus (HRV) serotypes 1-4 in Europe, we found that some strains could not be typed. However, when a monoclonal antibody directed to serotype 8 HRV was included in the typing assay, we detected seven 69 M-like (serotype 8) strains, six from Finland and one from Italy. The previously reported serotype 8 HRV strains, 69 M, B 37, and B 38 isolated in Indonesia, were of subgroup I specificity and presented a peculiar "super short" RNA electropherotype. In contrast, all the seven European strains possessed a long RNA pattern, and one of them had subgroup II specificity. Three of these strains were adapted to growth in cell cultures and were further characterized by neutralization and by Northern blot hybridization. They appeared to be closely related to serotype 8 HRV strain 69 M by neutralization, but showed partial homology with several human and animal strains by hybridization. The epidemiological importance of these serotype 8 strains circulating in Europe should be investigated, in view of their possible inclusion in a rotavirus vaccine.

Blotting, Northern↗
PubMed1993

Electropherotypes, serotypes, and subgroups of equine rotaviruses isolated in Japan.

Electropherotypes (ET), serotypes, and subgroups of equine rotaviruses isolated from foals in Japan were determined. The ETs of 136 isolates from 1981 through to 1991 were divided into six groups: ET-A-ET-F. The ET-A, -B, -C, -D, -E, and -F were present in 3, 1, 121, 9, 1, and 1 strains, respectively. Representative viruses of ET-A, -B, -C, and -D were identified as serotype G3. Viruses of ET-E and -F were identified as serotypes G 10 and G 5, respectively. The four representative viruses of serotype G 3 did not belong to either subgroup I or II. The two viruses of serotypes G 5 and G 10 belonged to subgroup I. Serotype G 3 strains possessing ET-C were prevalent among the foals throughout the study period.

Animals↗
PubMed1996

Modified latex agglutination test for rapid detection of Streptococcus pneumoniae and haemophilus influenzae in cerebrospinal fluid and direct serotyping of Streptococcus pneumoniae.

A modified latex agglutination test was designed and evaluated for the rapid detection of Streptococcus pneumoniae and haemophilus influenzae type b capsular antigens, and for direct serotyping of Streptococcus pneumoniae in the cerebrospinal fluid. Reagents were prepared by sensitizing latex particles with Omniserum (against 83 capsular serotypes of pneumococci) and Haemophilus influenzae type b burro antiserum. For serotyping reagents, latex particles were similarly coated with nine pneumococcal pool (a to I) antisera and 46 individual pneumococcal serogroup/serotype specific antisera. The test was performed on cerebrospinal fluid from 298 patients with suspected meningitis. Serotyping was done directly on untreated cerebrospinal fluid samples showing positive reactions with the Omniserum reagent. Pneumococcal or Haemophilus influenzae type b antigens were detected in 41 patients; in 32 of these the etiology was established by culture and in 2 by smear examination. Five of the remaining seven cases were judged clinically and by cytological examination of cerebrospinal fluid to have partially treated bacterial meningitis. In two cases the test was false positive. The overall sensitivity and specificity of the latex agglutination test for the detection of Streptococcus pneumoniae and Haemophilus influenzae type b antigens was 100% and 96.8% respectively. The commonest pneumococcal serotypes were type 1 (30%), types 6 and 19 (10% each). The latex agglutination test is rapid and simple to perform, yielding serotype data directly by testing of cerebrospinal fluid.

Adult↗
PubMed1996

Epidemiology of symptomatic human rotaviruses in Bangalore and Mysore, India, from 1988 to 1994 as determined by electropherotype, subgroup and serotype analysis.

Epidemiology of symptomatic rotaviruses from Bangalore and Mysore in Southern India was investigated. While serotype G3 predominated throughout the 7-year study period from 1988 to 1994 in Bangalore, serotype G1 was more predominant than serotype G3 in Mysore during 1993 and 1994. Serotype G2 strains were either not detected or infrequently observed in both the cities. However, several strains with subgroup I and 'short' RNA pattern that exhibited high reactivity with typing MAbs specific for serotype 2 as well as other serotypes were detected throughout the period. Among the nonserotypeable strains from both cities, several exhibited dual subgroup (SGI + II) or subgroup I specificity and 'long' RNA pattern indicating their probable animal origin. Notably, a gradual, yet highly significant reduction in rotavirus gastroenteritis, from 45.3% in 1988 to 1.8% during 1994, was observed in Bangalore in stark contrast to the consistently high (about 34%) incidence of asymptomatic infections among neonates by I321-like G10P11 type strains during the same period. Moreover, I321-like asymptomatic strains were not detected in children with diarrhea.

Antibodies, Monoclonal↗
PubMed1988

Protein patterns, serotyping and plasmid DNA profiles in the epidemiologic fingerprinting of Pseudomonas aeruginosa.

The epidemiological typing schemes based on serotyping, antibiotic susceptibility pattern, plasmid DNA profile, and protein patterns determined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) were evaluated for their usefulness in typing clinical isolates of Pseudomonas aeruginosa. The serotypability was lower (45%) than reported in the literature (85-95%). The most commonly found serotypes were O:1 (19%), O:11 (25%), O:6 (35%). The electrophoretic analysis of plasmid DNAs showed plasmids (molecular weight = 1 to greater than 40 megadaltons). Two plasmids of Mr 2 and 38 megadaltons were found in various serotypes. The restriction enzyme analysis of plasmid DNA showed identical DNA fragment patterns among distinct serotypes. The SDS-PAGE protein banding patterns of whole-cell proteins showed homogeneity among the strains. However, analysis of the soluble protein patterns of the strains showed sufficiently distinct protein profiles that can be used to differentiate between various strains. The results of this study demonstrate that the electrophoretic patterns of soluble proteins, in combination with plasmid DNA profile or serotyping, can be of value in the epidemiologic fingerprinting of clinical isolates of Pseudomonas aeruginosa.

Anti-Bacterial Agents↗
PubMed1985

Serotyping of Campylobacter species by combined use of two methods.

Five hundred strains of thermophilic Campylobacter spp. from sporadic cases of enteritis and from epidemiologically related infections connected with outbreaks were serotyped. The haemagglutination method of Penner and the slide agglutination method of Lior were used together. Greater discrimination was obtained by the use of two methods together than by either alone; 96% of sporadic strains were typed using a restricted set of typing sera. Of the sporadic strains, 50% fell within five Penner serotypes and 50% fell within four Lior serotypes, so the increased discrimination obtained by using both methods was particularly useful amongst these most common serotypes. In outbreaks associated with one serotype both methods gave consistent results, and in outbreaks due to multiple serotypes the two methods complimented each other.

Agglutination Tests↗
PubMed1983

Cross-reacting opsonic antibodies to clinically important pneumococcal serotypes after pneumococcal vaccination.

Opsonic activity of serum to pneumococcal serotypes 6 B, 9 V and 19 A was measured in 16 patients before and after immunization with a pneumococcal vaccine. The capsular polysaccharides of these serotypes are not included among, but are antigenically related to the vaccine polysaccharides. Patients responding to immunization with a twofold increase in serum antibodies to vaccine polysaccharides 6 A, 19 F and 23 F were studied. Increased opsonic activity towards serotypes 6 B, 9 V and 19 A was found in 12, four and ten patients respectively. In ten of the patients antibodies to serotypes 6 B, 9 V and 19 A were measured by a staphylococcal protein-A binding assay. A twofold increase in antibodies was found in postvaccination samples from ten, three and seven patients respectively. These results indicate that humans responding to pneumococcal vaccination, may also develop opsonic antibodies to other clinically important pneumococcal serotypes. The degree of cross-immunization appears to vary between individuals and between different pneumococcal serotypes.

Antibodies, Bacterial↗
PubMed1994

Relationship between fluconazole sensitivity of Candida albicans isolates from HIV positive patients and serotype, adherence and CD4+ lymphocyte count.

The sensitivity of fluconazole, serotype and adherence to buccal epithelial cells of 46 Candida albicans isolates was studied in vitro. Strains were isolated from 18 HIV-positive patients with variable CD4+ lymphocyte counts who were receiving long-term fluconazole therapy. Results showed that sensitivity to fluconazole correlated with serotype. Of the sensitive isolates 81.5% were serotype A and 18.5% serotype B, whereas of the resistant isolates 37.5% were serotype A and 62.5% serotype B. Sensitivity to fluconazole also correlated with adherence capacity, resistant isolates showing slightly less adherence to buccal epithelial cells than sensitive isolates. Weakly adherent isolates represented 87% of resistant isolates and only 63% of sensitive isolates. Resistant strains were isolated in highly immunocompromised patients. All strains isolated in patients with CD4+ lymphocyte counts above 200 lymphocytes/microliters were sensitive, whereas intermediate and resistant strains were isolated in patients with counts below 200 lymphocytes/microliters. Statistical analyses showed no correlation between adherence of isolates to buccal epithelial cells and CD4+ lymphocyte counts.

AIDS-Related Opportunistic Infections↗
PubMed1999

Extended serotyping scheme for Vibrio anguillarum with the definition and characterization of seven provisional O-serogroups.

The present paper summarizes the serotyping scheme of the fish pathogenic bacterium Vibrio anguillarum and defines seven additional O-serogroups. Strains, collected in our laboratories that were nontypable with antisera against the previously defined 16 O-serotypes, were used for generating new antisera and were characterized further by means of LPS profiles, Western blots, and serological reactions. On the basis of the results, it is suggested that the seven new O-serogroups are to be included in the existing serotyping system as serotypes O17-O23. However, the existence of further V. anguillarum strains that were not typable with any of the 23 O-antisera suggested the existence of additional O-serotypes within this species. The relevance of the description of additional O-serotypes for the species V. anguillarum is discussed.

Agglutination Tests↗
PubMed2005

Serotype distribution and mother-to-baby transmission rate of Streptococcus agalactiae among expectant mothers in Kuwait.

INTRODUCTION: Streptococcus agalactiae, also known as Group B Streptococcus (GBS), is a formidable pathogen that is commonly responsible for early-onset and late-onset infections with high morbidity and mortality in the neonatal period. Since this organism is usually acquired via the mother's birth canal during labor, this study investigated the maternal carriage rate, mother-to-baby transmission rate, and the common GBS serotypes found among expectant mothers and their babies in Kuwait. METHODS: The setting was the Maternity Hospital, Kuwait. Low vaginal-anorectal swabs (LVRS) and urine specimens were collected from 847 pregnant women during labor. Ear and umbilical swabs from their new-born babies were also collected. Each specimen was cultured on selective Todd-Hewitt media. Isolates were identified and serotyped by established methods. RESULTS: Of the 847 mothers, 124 (14.6%) were colonized and 74 (8.7%) babies were colonized, mainly at the umbilicus. The 124 GBS-positive mothers gave birth to 44 babies that were colonized by GBS at one or both sites, which corresponds to a mother-to-baby transmission rate of (35.5%). A total of 193 isolates were serotyped. The majority of the GBS isolates belonged to serotypes III (47; 24.3%), V (42; 21.8%), Ia (25; 12.9%), II and VI (15; 7.8%) each, and VII (11; 5.7%). Only 4 (2.1%) and 1 (0.5%) isolates belonged to serotypes Ib and IV respectively. No isolate belonged to serotype VIII and 33 (17.1%) were non-typable (NT).

Female↗