Occurrence of K1 and K2 serotypes and genotypic characteristics of extended spectrumβ-lactamases-producing Klebsiella pneumoniae isolated from selected hospitals in Malaysia

Objective:To determine the distribution,phenotypic and genetic background of extended spectrumβ-lactamases(ESBL)-producing Klebsiella(K.)pneumoniae clinical isolates associated with K1 and K2 serotypes in two selected hospitals in Malaysia.Methods:A total of 192 K.pneumoniae isolates were collected and subjected to antibiotic susceptibility,hypermucoviscosity test and multiplex PCR to detect the presence of K1-and K2-serotype associated genes.Multilocus sequence typing(MLST)was performed on ESBL-producing K.pneumoniae isolates presented with K1 and K2 serotypes,followed by phylogenetic analysis.Results:A total of 87 out of 192(45.3%)of the K.pneumoniae isolates collected were ESBL producers.However,only 8.3%(16/192)and 10.9%(21/192)of the total isolates were detected to carry K1-and K2-serotype associated genes,respectively.Statistical analysis showed that K1 and K2 capsular serotypes were not significantly associated with ESBL phenotype(P=0.196).However,they were significantly associated with hypervirulent,as demonstrated by the positive string test(P<0.001).MLST analysis revealed that ST23 as the predominant sequence type(ST)in the K1 serotype,while the ST in the K2 serotype is more diverse.Conclusions:Although the occurrence of ESBL-producing isolates among the hypervirulent strains was low,their coexistence warrants the need for continuous surveillance.MLST showed that these isolates were genetically heterogeneous.

Antibiotic Resistance Profile of Serotypes of Streptococcus pneumoniae Strains in Bangui, from 2017 to 2022: Case of Serotype 1

Goals: The aim of this study was to determine the antibiotic resistance profile of serotypes of Streptococcus pneumoniae strains circulating in Bangui. Methodology: A prospective and analytical analysis was carried out at the National Laboratory of Clinical Biology and Public Health from 2017 to 2022. The strains came from our study on the contribution to the study of antibiotic sensitivity of Streptococcus pneumoniae strains. The multiplex PCR test was used for its cost-effectiveness in terms of amplifiers which can be purified in order to be sequenced. It also makes it possible to detect several germs as well as their serotypes. For a PCR reaction, several elements are involved in the reaction medium or Master Mix. These are the desoxyribonucleotides (dNTPs), the magnesium ions (MgCl2) and the primers. A set of 14 primers divided into 3 classes were used. Class 1 primers served as an internal control by targeting the cpsA gene. It is a highly conserved gene found in capsular loci characterized to date. The primers of the second class were used to target specific serotypes by specific reactions (out of six possibilities). The group reaction was carried out using the primers of the third class in order to carry out an initial screening of the samples and to classify the pneumococcal isolates. Related serotypes were grouped based on the amplification of common genes. Using the technique of electrophoresis on agarose gel and an ultraviolet radiation device, the migration bands are then visualized and analyzed. The data collected had been entered into Excel 2010 and analyzed with Epi info 7. The exact Fischer chi2 test at the 5% threshold, the relative risk and its 95% confidence interval were used to compare the proportions and determine the associations. Results: 187 antibiotic-resistant strains of Streptococcus pneumoniae were collected. The average frequency of serotypes 1, 9A, 4 and untypeable identified were 43.59%, 18.18%, 18.27% and 39.57% respectively. The frequency of serotype 1 was predominant for the age group over five years old with 56.88%. The male sex was predominant with 55.08% for serotype 1. Resistance to penicillin and gentamicin for serotype 1 during this study, for the age group under 5 years old, was 77%. For serotypes 19A and 4, tetracycline resistance was predominant with 20% for the age group under 5 years. The resistance to penicillin and gentamicin of non-typeable serotypes was 33% for the age group under 5 years old. For the age group over 5 years old, resistance to erythromycin predominated at 37%. The distribution of serotypes by sex depending on antibiotic resistance was variable. There was a statistically significant association between identified serotypes and antibiotic resistance (p Conclusion: The study determined serotypes 1, serotypes 19A, serotypes 4 and non-typeable serotypes. These results would be due to the quality of vaccination or poor protection of vaccines.

Prevalence of Precancerous Lesions Based on Digital Cervicography with VIA/VILI among Women Positive for High-Risk Human Papillomavirus Serotypes: A Screening Center-Based Study in Cameroon

Background: Since 2021, high-risk Human Papilloma Virus (HR-HPV) testing has been the recommended screening test for cervical cancer for all settings;either used alone in a “test and treat” strategy, or with a triage test, with or without biopsy, before treatment. Cameroon has rolled out immunization against HPV 16 and 18, but studies show a higher prevalence of non-16/18 HR-HPV types. Objectives: Determine the prevalence of precancerous lesions, in women with HR-HPV infection and evaluate association of digital cervicography (DC) VIA/VILI positivity with HPV serotype, as a measure of their contribution to precancer and cancer incidence. Methodology: The study was cross-sectional, descriptive, and analytic. It took place at the Etoug-Ebe and Ekoudoum Baptist Hospitals in Yaoundé, during the period April-September 2022. We reviewed the records of women screened for cervical cancer between February 2020 and December 2021 and evaluated the prevalence of lesions on digital cervicography (DC) with VIA/VILI for women positive for HR-HPV serotypes. The data were analyzed using SPSS version 20.0 for Windows. P values Results: We identified 315 cases with a positive HR-HPV deoxyribonucleic acid (DNA) test, 224 (71.1%) had a DC VIA/VILI triage test done. Of these, 30 (13.4%) women had a positive DC VIA/VILI, with five women (2.2%) having lesions suggestive of cancer. Out of 11 cases positive for HPV 16 alone, 05 (45.5%) had a positive DC VIA/VILI test. Of the 14 cases positive for HPV 18 alone, 03 (21.4%) had a positive VIA/VILI, meanwhile only 19 (10.7%) of the 177 cases positive for non-16/18 HPV had a positive VIA/VILI test. Conclusion: A high proportion of women (13.4%) with HR HPV had a positive DC VIA/VILI, with a significant proportion (2.2%) having lesions suggestive of invasive cervical cancer HR-HPV serotype was associated with DC VIA/VILI positivity;HPV 16 had the strongest association (45.5%), followed by HPV 18 (21.4%), and non-16/18 HR-HPV (10.7%), suggesting a decreasing order of oncogenicity.

Changes in the Rate of Human Papilloma Virus Serotypes after Vaccine Implementation: A Descriptive Study

Background: The main objective of this study is to describe the rate of the different serotypes of HPV in cervical cytologies and biopsies in three different periods: 2002-2006 (prior to the implementation of the vaccination programs in Spain), 2009-2011 (shortly after this implementation) and 2020 (almost 15 years after introduction of the vaccine) at a single hospital. Methods: This is an observational, descriptive, retrospective study based on the review of the results of the determination of the HPV serotype using the commercial kit (Genomica®;PharmaMar LTD) in cervical liquid-based cytologies and biopsies at a single large tertiary hospital, Hospital Clínico San Carlos, in Madrid, Spain. We have collected the data from three different time periods: 2002-2006;2009-2011, and 2020 to try to understand the potential changes associated with the use of the vaccine. Results: In these time periods we have reviewed the data from 1420 women. In the three periods the most frequent serotype was HPV 16, followed by HPV 18 or a combination of both. The most frequent low risk serotype was HPV 6 followed by the combination of HPV 6 and 11. It has been verified in our study that the prevalence of the category “others”, constituted by the three risk groups, has undergone a progressive increase, beginning with an infection rate of 65.43% in 2002-2006 to finally rise up to 90.92% in the year 2020. Conclusions: Our study reveals an increase in the number of infections by the HPV serotypes that are not included in the tetravalent vaccine.

Membrane vesicles derived from Streptococcus suis serotype 2 induce cell pyroptosis in endothelial cells via the NLRP3/Caspase-1/GSDMD pathway

Streptococcus suis serotype 2(S.suis 2)is a zoonotic pathogen that clinically causes severe swine and human infections(such as meningitis,endocarditis,and septicemia).In order to cause widespread diseases in different organs,S.suis 2 must colonize the host,break the blood barrier,and cause exaggerated inflammation.In the last few years,most studies have focused on a single virulence factor and its influences on the host.Membrane vesicles(MVs)can be actively secreted into the extracellular environment contributing to bacteria-host interactions.Gram-negative bacteria-derived outer membrane vesicles(OMVs)were recently shown to activate host Caspase-11-mediated non-canonical inflammasome pathway via deliverance of OMV-bound lipopolysaccharide(LPS),causing host cell pyroptosis.However,little is known about the effect of the MVs from S.suis 2(Gram-positive bacteria without LPS)on cell pyroptosis.Thus,we investigated the molecular mechanism by which S.suis 2 MVs participate in endothelial cell pyroptosis.In this study,we used proteomics,electron scanning microscopy,fluorescence microscope,Western blotting,and bioassays,to investigate the MVs secreted by S.suis 2.First,we demonstrated that S.suis 2 secreted MVs with an average diameter of 72.04 nm,and 200 proteins in MVs were identified.Then,we showed that MVs were transported to cells via mainly dynamin-dependent endocytosis.The S.suis 2 MVs activated NLRP3/Caspase-1/GSDMD canonical inflammasome signaling pathway,resulting in cell pyroptosis,but it did not activate the Caspase-4/-5 pathway.More importantly,endothelial cells produce large amounts of reactive oxygen species(ROS)and lost their mitochondrial membrane potential under induction by S.suis 2 MVs.The results in this study suggest for the first time that MVs from S.suis 2 were internalized by endothelial cells via mainly dynamin-dependent endocytosis and might promote NLRP3/Caspase-1/GSDMD pathway by mitochondrial damage,which produced mtDNA and ROS under induction,leading to the pyroptosis of endothelial cells.

Echovirus serotypes circulating in Malaysia from 2002 to 2013

Objective:To identify the circulating serotypes of human echovinis in Malaysia from 2002 in 2013.Methods:A toial of 31 retrospective samples from non-polio acute flacid paralysis,hand-food-and-mouth disease,viral meningitis and enterovirus cases were subjected to amplification of partial VPI gene by RT-PCR.Results:Sequencing and phylogeneiic analysis of the partial sequences identified presence of human echovinis and human coxsackie viruses.It was found that echovinis 11 was the commonly circulating serotype followed by echovinis6.echovinis 7.echovinis 3.echovinis 9.echovinis 30 and echovinis I in decreasing order.Additionally two types of human coxsackie virus isolates were detected which were coxsackie A24 and B3.Condusions:From the findings,there is a possibility that echovinis 11 is the predominant serotype among Malaysian patients with echovinis infection.However,a larger sample size will yield a more confident result to support this evidence.____________________

Spatial Trend of Foot and Mouth Disease Virus (FMDV) Serotypes in Cattle and Buffaloes, Pakistan

The present study describes the frequency of Foot and Mouth Disease （FMD） virus serotypes （O, A and Asia-l） in major regions （all provinces） of Pakistan using Indirect Sandwich ELISA. Also, spatial distribution of various FMD serotypes and their comparison is discussed. A total of 590 samples （Epithelial tissue） have been analyzed during a period of five years （2005-2009）. Out of 590 samples, 180 were found positive, giving an overall confirmation of FMDV about 33.2 %. Of the prevalent serotypes, FMDV ＇O＇ serotype caused most outbreaks （20.7 %）, followed by serotype A （6.6 %） and serotype Asia-1 （4.6 %） while there was no positive case of type ＇C＇. The study clearly showed that the disease was more frequent in the agro-climatic zones than in hilly areas. Based on the data of 590 samples （〉50 outbreaks）, the overall prevalence of FMDV in cattle and buffaloes in Pakistan was 33.2 %, while in cattle alone, it was 37.1%, higher than in buffalo （28.7 %）. There were eight cases of mixed serotypes infection, indicating the presence of endemic state of disease. Another significant feature was the change over time. In phase-I （2005-2007）, there was an overall prevalence of 29.4 %, while the occurrence of the serotype O, A and Asia-1 was 20.4 %, 2.9 % and 4.7 %, respectively. During phase-II （2008-2009）, the overall prevalence was 59.21%, while those of serotype O, A and Asia-1 were 22.4 %, 31.6 % and 4.0 %, respectively. This clearly indicated a shift from serotype O to A, which may help to explain the occurrence of more severe outbreaks, despite vaccination.

Loop-Mediated Isothermal Amplification (LAMP) for the Detection of <i>Listeria monocytogenes</i>and Major Pathogenic Serotypes

Rapid identification and characterization of Listeria monocytogenes are required for the food industry, epidemiological studies, and disease prevention and control. However, typing procedures are labor-intensive and time-consuming, and they require technical expertise, a panel of sera and reference culture strains or sophisticated and expensive equipment. To improve upon traditional diagnostic methods for L. monocytogenes we developed and evaluated an efficient procedure for the specific identification of L. monocytogenes and the major pathogenic serotypes of the species based on loop-mediated isothermal amplification (LAMP). Four individual reactions were designed using primers targeting any L. monocytogenes serotypes (LAMP-AS) and the 1/2a (LAMP-1/2a), 1/2b (LAMP-1/2b), and 4b (LAMP-4b) serotypes. The procedure distinguished L. monocytogenes from closely genetically related species and the targeted serotypes. Cross-reactivity with a few rare serotypes isolated from food or clinical samples did not impair the usefulness of the procedure. Thus, our approach constitutes a fast, easy and low-cost alternative for L. monocytogenes diagnosis and serotyping and may be useful for surveillance and epidemiological investigation programs.

Serotypes of Non-O157 Shigatoxigenic <i>Escherichia coli</i>(STEC)

Non-O157 STEC has been shown to have a diverse ecological distribution among food-animals. It has been associated with both outbreaks and individual cases of severe illness. This group of the organisms is now considered as a major contributor to human disease. The clinical description of the diseases caused by these organisms is reviewed. The host specificity of these pathogens is described and discussed. These organisms appear widespread among food animals like cattle and sheep, and can therefore affect a range of foods directly from the meat and excretions of these animals being used in farming practices. This article reviews the origins, diversity and pathogenesis of non-O157 STEC.

Serotypes of Bacteria Encountered in Childhood Purulent Meningitis in Children in Parakou (Benin) in 2011

Introduction: In the North-Benin, there are three agents causing pediatric purulent meningitis outside the neonatal period. These are: Streptococcus pneumoniae, Neisseria meningitidis and Haemophilus influenzae type b. The aim of this research work was to investigate bacteria serotypes that caused childhood purulent meningitis in the pediatric unit of the Borgou à Regional University Teaching Hospital (CHUD-Borgou) located in Parakou (North-Benin). Patients and Methods: Through a prospective and descriptive study centered on children aged 0 to 5 years old suspected of meningitis and hospitalized, the cerebrospinal fluid (CSF) samples of those children were analyzed at the WHO reference laboratory in Banjul for serotyping by real time polymerase chain reaction (RT/PCR). Results: Among the 1396 children hospitalized during that period, 366 were suspected of meningitis and had benefitted from lumbar puncture. Among those 366 suspected cases, 51 cases of purulent meningitis were confirmed after CSF cytobacteriological and biochemical test at the CHUD-Borgou laboratory. Among 51 CSF samples in which purulent meningitis was confirmed, 44 were sent to Banjul. In addition, 310 CSF samples from non-confirmed cases of meningitis were also sent to Banjul. In the whole set of samples sent for real time PCR, 151 cases of Streptococcus pneumoniae (42.7%) were found, 5 cases of Neisseria meningitidis (1.4%) and 1 case of Haemophilus influenzae (0.3%) were also encountered. As regards Streptococcus pneumonia, the serotypes encountered were: 1, 3, 4, 5, 7F, 8, 9V, 9V/9A, 9N/9L, 14, 18C, 19A, 23F, 33F as well as non typed and non typable serotypes. As for Neisseria meningitidis, only serogroup A was found in it. For Haemophilus influenzae, only serotype b was identified. Conclusion: Four non vaccine serotypes (8, 9V/9A, 9N/9L and 33F), non typed and non typable serotypes which are not covered by 13-valent pneumococcal conjugate vaccine (PCV 13) were identified. This highlights the need to enhance surveillance of pediatric purulent meningitis and serotyping by RT/PCR of all CSF samples in order to adapt if necessary future new pneumococcal vaccines to circulating non vaccine serotypes.

Study on Serotypes and Auxotypes of Neisseria Gonorrhoeae in Guangzhou

Objective:To investigate the serotypes and auxotypes distribution of Neisseria gonorrhoeae in Guangzhou. Method: 131 strains of Neisseria gonorrhoeae wereserotyped by co-agglutination test and 108 strains wereauxotyped by La Scolea's method. Results: Out of 131 strains of Neisseria gonorrhoeae,87.8% (115/131) were WⅡ/WⅢ, while 9.9% (13/131) wereWⅠ. The most important auxotypes were Proto, Pro and ILe,42.6% (46/108), 21.3% (23/108) and 12.0%, respectively. WⅡ/WⅢ was distributed among the all auxotypes aboveand WI found only in both Proto and Pro. Conclusion: The study illustrated the prevailing serotype,WⅡ/WⅢ, and higher prevalence of Ile- in Guangzhou.

Comparative Assessment of Growth and Gene Regulation between 4 Serotypes of <i>Streptococcus pneumoniae</i>in Broth and Cell Culture

Streptococcus pneumoniae is a medically important pathogen capable of causing human infections of pneumonia, bacteremia, otitis media, and meningitis. Although there are vaccinations available, infections with S. pneumoniae still remains a global problem. S. pneumoniae is a highly adaptable bacterial species with numerous serotypes based on capsular polysaccharides. The different serotypes vary in their ability to colonize and causing pathology. Here we compared the regulation of five different virulence factors from four common serotypes of S. pneumoniae that vary in their carriage, morbidity, and mortality rates in the human population using two different in vitro methods, broth and cell culture. We determine that there is variation of virulence factor gene regulation within a serotype using two different culture methods, and variation between the serotypes in the same culture condition. The regulation of genes appeared to have a correlation with the ability of the various serotypes to grow in broth culture, adhere to cultured lung cells, and invade the cultured lung cells, as serotypes that shared similar regulation of virulence factors tended to behave similarly in culture. Many studies with S. pneumoniae rely on the use of one selected serotype, but since there is a wide variation in the growth and regulatory mechanisms of these bacteria. As demonstrated here, future studies should utilize more strains in models before concluding mechanisms of pathobiology.

Enzyme-Linked Immunosorbent Assay for Pasteurella Multocida in Rabbits and Serotypes of the Isolates

The control and prevention of pasteurellosis in rabbits which makes the hosts unsuitable for experimental use raised the needs to improve and simplify the procedures of Enzyme-Linked Immunosorbent Assay (ELISA) for detect of antibody against P.multocida. A comparison on the sensitivity and specificity of bacterial culture of antemortem and postmortem samples, complement fixation test and enzyme-linked immunosorbent assay of 11 apparently healthy adult rabbits was conducted. The incidence rates showed 45.45%,54.54% and 72.73% respectively. The sensitivity for the three methods were 0.63, 0.67,and 1.00,and specificity for them were 1.00, 0.67 and 1.00 respectively. Somatic serotypes of isolates of P.multocida from rabbits of three groups (rabbits of group 2 were with clinic signs, those of groups 1 and 3 were apparently healthy) revealed no remarkable differences,and the predominant types were type 3 and type 3, 4. This was somewhat different from the reports derived from other states. As the antigen of different serotype used in ELISA may have different sensitivity and specificity, which is affected also by different preparation method, a type non-specific antigen should be selected to meet such request. The trial of accomplishment of ELISA without positive and negative controls was presented for discussion.

Antimicrobial susceptibility and serotypes of Neisseria meningitidis and Streptococcus pneumoniae in Sri Lanka:Experience from the National Reference Laboratory

Objective:To determine the antimicrobial susceptibility and serotypes of Neisseria(N.)meningitidis and Streptococcus(S.)pneumoniae in Sri Lankan patients.Methods:We retrospectively analyzed 11 blood culture specimens from suspected patients with invasive meningococcal disease and 26 S.pneumoniae clinical isolates.We tested 6 antimicrobials against N.meningitidis and 12 antimicrobials against S.pneumoniae.Meningococcal serogroup was determined by realtime PCR and Quellung serotyping was used for pneumococcal analysis.Results:N.meningitidis serogroup B was the most common in this study.Intermediate-susceptibility to penicillin was seen in 75.0%(6/8)of strains.Susceptibility to ciprofloxacin,levofloxacin and cotrimoxazole was 62.5%(5/8),62.5%(5/8)and 87.5%(7/8),respectively.Excellent susceptibility was seen in cefotaxime and meropenem.In S.pneumoniae,the most common serotype was 19F in both invasive and non-invasive pneumococcal diseases.The majority of strains showed multidrug resistance.Penicillin non-susceptibility in non-meningeal strains were 13.6%and all meningeal strains were penicillin resistant.Erythromycin was highly resistant in both groups.Amoxicillin showed excellent susceptibility in non-invasive pneumococcal diseases strains.Linezolid,levofloxacin and vancomycin showed 100.0%susceptibility in all pneumococcal isolates.Conclusions:Implementation of vaccines should be considered,especially for children and high-risk populations.This may contribute to reducing pneumococcal and meningococcal invasive disease burden and help prevent emergence of antimicrobial resistant strains.

Serotypes and Antimicrobial Susceptibility Patterns of Nasopharyngeal Pneumococci Isolated from HIV-Infected Children in Selected Pediatric Clinics in Nairobi, Kenya

Background: Streptococcus pneumoniae (pneumococcus) is one of the most frequent causes of bacterial infection in children and is a leading cause of otitis, sinusitis, pneumonia, and meningitis worldwide. Nasopharyngeal colonization is a risk factor for pneumococcal disease, a leading cause of complications and death in infants. HIV-infected persons are at high risk of invasive pneumococcal disease. Method: Nasopharyngeal swabs were collected from 296 HIV infected children below five years recruited from Gertrude’s Children hospital and Nazareth Hospital Nairobi, Kenya. The nasopharyngeal swabs were processed to isolate S. pneumoniae, which were serotyped and tested for drug susceptibility. Results: The carriage prevalence of S. pneumoniae in the study was 30.4% while the isolated serotypes were (in order of decreasing frequency): 35B, 19F, 3, 13, 15A, 11A, 16F, 7C and 23A. Most of the serotypes were resistant to the commonly used antibiotics but all were susceptible to vancomycin and chloramphenicol. Conclusion: Carriage prevalence of nasopharyngeal S. pneumonia in HIV infected children was lower than that of similar prevalence studies in children. Most of the S. pneumoniae isolates were however non pneumococcal vaccine isolates.

A Cross Sectional Sero-Study of Verocytotoxigenic Escherichia coli (VTEC) Serotypes in Apparently Healthy and Diarrhoeic Cattle in Abuja, Federal Capital Territory (FCT), Nigeria

It is reckoned worldwide that verocytotoxigenic Escherichia coli (VTEC) serotypes are important food borne pathogens causing severe health problems in humans. A cross sectional epidemiological study was carried out to determine the prevalence of VTEC serotypes (O157 and non O157) in both apparently healthy and diarrhoeic cattle in Abuja, FCT. A total of 718 faecal samples collected from abattoirs and cattle herds from Abuja, FCT representing 381 from apparently healthy and 337 from diarrhoeic cattle were analyzed. Primary isolation of typical E. coli was done using Eosin Methylene Blue (EMB) agar and performing biochemical tests. Samples were further analyzed using Cefixime, Tellurite-Sorbitol McConkey (CT-SMAC) agar to identify sorbitol and non sorbitol fermenting E. coli. Further characterization of both the sorbitol fermenting and non sorbitol fermenting E. coli was done using commercially procured latex agglutination test kits from Oxoid, United Kingdom. The prevalence of VTEC O157 in apparently healthy cattle was 1.84% and 2.96% for diarrhoeic cattle while the prevalence of non O157 VTEC was 3.67% and 7.12% for apparently healthy and diarrhoeic cattle respectively. There was no strong association (p > 0.05) between faecal consistency and infection with VTEC O157. A strong association (p < 0.05) however existed between faecal consistency and infection with non-O157 VTEC. Diarrhoeic cattle appear likely to be more affected. The implication of the study is that individuals in contact with cattle such as veterinarians, abattoir workers and cattle herdsmen are at risk of exposure to VTEC and proper hygienic control measures should be adopted.

Biovars and Serotypes of Ureaplasma Urealyticum among Chinese Women Undergoing Routine Gynecologic Exam, Women with Sexually Transmitted Infections, and Female Sex Workers

Objectives: To characterize the distribution pattern of biovars and scrotypes or Ureaplasma urealyyicum in normalhealthy women, sexually transmitted infections clinic clients,and in sex workers. Methods: We cultured cervical swabs taken from 261physical check-up clients, 599 STI clinic outpatients and 98 sexworkers using commercial selective medium. Some positivecultures were further biotyped and serotyped by PCR. Results: (1) U. urealyticum is more commonly isolated in sexworkers (90.8%) than in the physical check-up group (60.9%)or the STI outpatient group (61.3%) (P<0.001). (2) Biovar 1of U. 'realyticum (95.0%), especially single infection ofserotype 1. 3, and 6 of biovar 1, is commonly found in healthywomen. (3) Biovar 2 infection of U urealyticum is moreprevalent in sex workers (28.1%) and STI outpatients group(26.6%) than that in the physical check-up group (4.9%) (P<0.001). (4) Mixed infection caused by more than one serotypeof U urealyticum increased from physical check-up group(8.6%) to STI utpatients (12.4%) to sex workers (23.9%) (P<0.01). (5) There is no statistically significant difference in thedistribution of serotype 1, 3, and 6 of biovar 1 among thesethree groups (P=0.763). (6) The PCR method described here isrelatively simple, rapid and specific for the biotyping andserotyping of biovar 1 of U urealyticum. Conclusion: We should pay more attention to biovar 2 andmixed infections of U. urealyticum than single infection ofhiovar 1 in clinic practice. PCR is a good method for biotypingand serotvping.

Investigation on Biovars and Serotypes of Ureaplasma Urealyticum in Cervix in Chinese Gynecologic Check-up Population, Sexually Transmitted Infections Clinic Clients and Sex Workers

Objectives: To characterize the distribution pattern biovars and serotypes of Ureaplasma urealyticum in normahealthy women, sexually transmitted infections clinic clienand in sex workers. Methods: We cultured cervical swabs taken from 26physical check-up clients, 599 STI clinic outpatients and 9sex workers using commercial selective medium. Sompositive cultures were further biotyped and serotyped bPCR. Results: (1) Biovar 1 of U urealyticum (95.0%), especiallsingle infection of serotype 1, 3, and 6 of biovar 1,commonly found in healthy women. (2) U urealyticummore commonly isolated in sex workers (90.8%) than iphysical check-up group (60.9%) and STI outpatients grou(61.3%) (P<0.001). (3) Biovar 2 infection of U urealyticuris more prevalent in sex workers (28.1%) and SToutpatients group (26.6%) than that in physical check-ugroup (4.9%) (P<0.001). (4) Mixed infection caused bmore than one serotype of U. urealyticum is increasing fromphysical check-up group (8.6%) to STI outpatients (12.4%and to sex wokers (23.9%) (P<0.01). (5) There is nstatistic difference in the distribution of serotype 1, 3, and of biovar 1 among these three groups (P=0.763). (6) ThPCR method described here is relatively simple, rapid anspecific for the biotyping and serotyping of biovar 1 of Uurealyticum. Conclusion: we should pay more attention to biovarand mixed infection than single infection of biovar 1 of Uurealyticum in clinic practice. PCR is a good method ibiotyping and serotyping.

A Simple Procedure for Extraction of Surface Protein of Salmonella Serotypes and Escherichia coli Strains Isolated from Poultry and Pigs

Salmonella and E.coli possess different surface protein structures that can induce protective immune responses.Identification of these proteins capacitates development of diverse applications in prevention and diagnosis that contribute to effectively control disease-causing enterobacteria pathogens such as Salmonella and E.coli.A simple procedure for obtaining protein complexes of Salmonella serotypes and E.coli is performed in this study.A sonication process with heat treatment of whole bacteria induced the release of protein complexes.Concentration of the protein extract was quantified using protein quantification Kits-Rapid,and protein complex profile was obtained by SDS-PAGE(Sodium dodecyl sulfate polyacrylamide gel electrophoresis)and silver staining.The concentrations of protein ranged from 29.45 to 45.35μg/mL in the Salmonella protein extracts,and from 25.35 to 36.72μg/mL in the E.coli protein extracts.Six major groups of proteins from E.coli(YfiO,NipB,OmpF,YfgL,Talc,YaeT)and four major groups of proteins from Salmonella(Flagellin,OmpA,Porin,SEF21)were preliminarily determined by a simple procedure of extraction based on the molecular weight.

Establishment and Modification of Ninety-seven Pneumococcal Serotyping Assays Based on Quantitative Real-time Polymerase Chain Reaction

Objective To establish and modify quantitative real-time polymerase chain reaction(qPCR)-based serotyping assays to distinguish 97 pneumococcal serotypes.Methods A database of capsular polysaccharide(cps)loci sequences was generated,covering 97 pneumococcal serotypes.Bioinformatics analyses were performed to identify the cps loci structure and target genes related to different pneumococcal serotypes with specific SNPs.A total of 27 novel qPCR serotyping assay primers and probes were established based on qPCR,while 27 recombinant plasmids containing serotype-specific DNA sequence fragments were constructed as reference target sequences to examine the specificity and sensitivity of the qPCR assay.A panel of pneumococcal reference strains was employed to evaluate the capability of pneumococcal serotyping.Results A total of 97 pneumococcal serotyping assays based on qPCR were established and modified,which included 64 serotypes previously reported as well as an additional 33 serotypes.Twenty-seven novel qPCR serotyping target sequences were implemented in the pneumococcal qPCR serotyping system.A total of 97 pneumococcal serotypes,which included 52 individual serotypes and 45 serotypes belonging to 20 serogroups,could not be identified as individual serotypes.The sensitivity of qPCR assays based on 27 target sequences was 1–100 copies/μL.The specificity of the qPCR assays was 100%,which were tested by a panel of 90 serotypes of the pneumococcal reference strains.Conclusion A total of 27 novel qPCR assays were established and modified to analyze 97pneumococcal serotypes.