Australian Meningococcal Surveillance Programme annual report, 2019.

Invasive meningococcal disease (IMD) is a notifiable disease in Australia, and both probable and laboratory-confirmed cases of IMD are reported to the National Notifiable Diseases Surveillance System (NNDSS). In 2019, there were 206 notifications of IMD. Of these, 202 were laboratory-confirmed cases analysed by the reference laboratories of the Australian National Neisseria Network (NNN). Of the 202 laboratory-confirmed cases of IMD, 167 were confirmed by bacterial culture and 35 by nucleic acid amplification testing, and all had the serogroup determined. Fine typing was available on 146 samples (146/202, 72%). Neisseria meningitidis serogroup B (MenB) infections accounted for 50.0% (101/202); MenW for 26.2% (53/202); MenY for 20.8% (42/202) and MenC for 3.0% of cases (6/202). Of the MenW cases, 88% were PorA antigen type P1.5,2, and 65% of these (24/37) were sequence type 11, the hypervirulent strain reported in recent outbreaks in Australia and overseas. The primary peaks of IMD notifications in Australia in 2019 were observed in infants less than 1 year of age (36/202, 18%) and in adults aged 65 years or older (39/202, 19%). MenB infections predominated in those aged less than 5 years and those aged 15-19 years, whereas MenW and MenY infections predominated in those aged 45 years or more. All 167 IMD isolates were tested for antimicrobial susceptibility. One isolate out of these 167 (0.6%) was resistant to penicillin with an MIC ≥ 1mg/L; 154/167 isolates (92%) had decreased susceptibility to penicillin. All isolates were susceptible to ceftriaxone and ciprofloxacin, and one isolate was resistant to rifampicin.

Lancefield Whole Blood Killing Assay to Evaluate Vaccine Efficacy.

While the Lancefield whole blood killing assay is named after the renowned streptococcal researcher Rebecca Lancefield, the protocol was first described by Todd in 1927 (Br J Exp Pathol 8:1-5, 1927). Initially, the assay was used to identify novel Group A Streptococcal (GAS) serotypes through the supplementation of non-immune human blood (often from infants) with type-specific antisera prepared in rabbits (Lancefield, J Exp Med 106:525-544, 1957; Maxted, Br J Exp Pathol 37:415-422, 1956) and to demonstrate the impressive longevity of type-specific immunity in patients following invasive GAS infection (Lancefield, J Exp Med 110:271-292, 1959). The modern assay is routinely used to screen defined GAS mutants (Wessels, Bronze, Proc Natl Acad Sci U S A 91:12238-12242, 1994; Zinkernagel et al., Cell Host Microbe 4:170-178, 2008) or transposon libraries (Le Breton et al., Infect Immun 81:862-875, 2013) for enhanced susceptibility to opsonophagocytic killing or to screen vaccine antisera (Salehi et al., mSphere 3:e00617-e00618, 2018) or other serological preparations (Reglinski et al., Sci Rep 5:15825, 2015) for anti-streptococcal activity.

Reduction of antimicrobial resistant pneumococci seven years after introduction of pneumococcal vaccine in Iceland.

BACKGROUND: Penicillin non-susceptible (PNSP) and multi-resistant pneumococci have been prevalent in Iceland since early nineties, mainly causing problems in treatment of acute otitis media. The 10-valent protein conjugated pneumococcal vaccine (PHiD-CV) was introduced into the childhood vaccination program in 2011. The aim of the study was to investigate the changes in antimicrobial susceptibility and serotype distribution of penicillin non-susceptible pneumococci (PNSP) in Iceland 2011-2017. METHODS AND FINDINGS: All pneumococcal isolates identified at the Landspítali University Hospital in 2011-2017, excluding isolates from the nasopharynx and throat were studied. Susceptibility testing was done according to the EUCAST guidelines using disk diffusion with chloramphenicol, erythromycin, clindamycin, tetracycline, trimethoprim/sulfamethoxazole and oxacillin for PNSP screening. Penicillin and ceftriaxone minimum inhibitory concentrations (MIC) were measured for oxacillin resistant isolates using the E-test. Serotyping was done using latex agglutination and/or multiplex PCR. The total number of pneumococcal isolates that met the study criteria was 1,706, of which 516 (30.2%) were PNSP, and declining with time. PNSP isolates of PHiD-CV vaccine serotypes (VT) were 362/516 (70.2%) declining with time, 132/143 (92.3%) in 2011 and 17/54 (31.5%) in 2017. PNSP were most commonly of serotype 19F, 317/516 isolates declining with time, 124/143 in 2011 and 15/54 in 2017. Their number decreased in all age groups, but mainly in the youngest children. PNSP isolates of non PHiD-CV vaccine serotypes (NVT) were 154/516, increasing with time, 11/14, in 2011 and 37/54 in 2017. The most common emerging NVTs in 2011 and 2017 were 6C, 1/143 and 10/54 respectively. CONCLUSIONS: PNSP of VTs have virtually disappeared from children with pneumococcal diseases after the initiation of pneumococcal vaccination in Iceland and a clear herd effect was observed. This was mainly driven by a decrease of PNSP isolates belonging to a serotype 19F multi-resistant lineage. However, emerging multi-resistant NVT isolates are of concern.

Cephalotaxine inhibits Zika infection by impeding viral replication and stability.

The Zika virus (ZIKV) is a mosquito-borne flavivirus that has reemerged as a serious public health problem around the world. Syndromes of infected people range from asymptomatic infections to severe neurological disorders, such as Guillain-Barré syndrome and microcephaly. Screening anti-ZIKV drugs derived from Chinese medicinal herbs is one method of identifying antiviral agents. In this paper, we report that (1) Cephalotaxine (CET), an alkaloid isolated from Cephalotaxus drupacea, was effective in inhibiting ZIKV activity in vitro (i.e., in Vero and A549 cell lines) and (2) the mechanisms which underlie these effects involve virucidal activity and a decrease in viral replication. Specifically, CET was found to decrease ZIKV RNA and viral protein expression, inhibit ZIKV replication, and inhibit ZIKV mRNA/protein production. We also determined that CET is effective in inhibiting dengue virus 1-4 (DENV1-4). Taken together, our findings indicate that CET could be an effective lead compound in the treatment of ZIKV and also suggest that further investigation and development of CET-derived drugs may lead to a new class of anti-Flavivirus medications.

Epidemiological and molecular characterization of invasive Streptococcus pneumoniae isolated following introduction of 7-valent conjugate vaccine in Kinki region, Japan, 2008-2013.

Streptococcus pneumoniae is one of the most common bacteria causing community-acquired pneumonia and meningitis. The use of 7-valent pneumococcal conjugate vaccine (PCV7) has reduced the incidence of pneumococcal disease while changing pneumococcal population through herd immunity and non-vaccine pneumococci replacement. This study investigated molecular epidemiologic characteristics of pneumococcal strains in the Kinki region of Japan from 2008 to 2013. A total of 159 invasive pneumococcal isolates were characterized by serotyping, antibiotic susceptibility testing, PCR analysis of penicillin-binding protein genes, multilocus sequence typing (MLST), and pulsed-field gel electrophoresis (PFGE). In adult populations, pediatric PCV7 introduction decreased isolates expressing PCV7 serotypes via herd immunity and increased isolates expressing non-PCV7 serotypes. The rate of penicillin resistance and isolates with alterations in all three pbp genes was higher in PCV7 type isolates than in non-PCV7 type isolates. In MLST analysis, all of serotype 19F isolates were of the same sequence type, ST236, which is the antimicrobial-resistant clone Taiwan19F-14, and the majority of serotypes 23F and 19A isolates were of ST1437 and ST3111 respectively, which are the predominant clones of antimicrobial-resistant pneumococci in Japan. In PFGE profiles, serotype 6B-ST2224, serotype 19F-ST236, serotype 19A-ST3111, and serotype 23F-ST1437 formed six separate clusters composed of genetically identical strains, and genetically identical serotype 22F-ST433 formed two different clusters between the pre- and post-PCV7 period. The results of molecular analysis suggest the spread and persistence of these identical antimicrobial resistant clones in the Kinki region and genetic changes of epidemic clone serotype 22F-ST433 before and after pediatric PCV7 introduction.

Phenotypic and genotypic characterization of serogroup 6 Streptococcus pneumoniae isolates collected during 10-valent pneumococcal conjugate vaccine era in Bulgaria.

Serogroup 6 remains common in the pneumococcal-conjugated vaccine era in Bulgaria; therefore, we investigated its clonal and serotype dynamics. The antibiotic susceptibilities were assessed by broth microdilution. Strains identified as serogroup 6 with latex agglutination method were subjected to serotype-specific PCRs. Erythromycin-resistant strains were analyzed by PCR for presence of ermB and mefE genes. MLST was performed to define clonal composition of the sequence types (STs). Serogroup 6 was represented by 40 (13.3%) from 301 invasive and non-invasive Streptococcus pneumoniae isolates. Molecular serotyping revealed new emerging serotype 6C (6.6%), not detected in pre-vaccine era. Among unvaccinated patients, mostly we observed serotypes 6А (57.1%) and 6В (28.6%). Serotype 6C was distinctive for vaccinated children (64%), followed by 6A (24%). Penicillin and ceftriaxone non-susceptible serogroup 6 strains were 65% and 5%, respectively; erythromycin- and clindamycin-resistant were 70.0% and 52.5%, respectively. Multidrug-resistant strains were 57.5%. Prevalent genetic determinant for macrolide resistance was ermB gene (75%). MLST revealed 17 STs into 5 clonal complexes and 7 singletons. Predominant genetic lineage was CC386, represented by MDR-6C non-invasive strains. Serotype 6B, principally responsible for invasive diseases in the pre-vaccine era, retreated this position to serotype 6A.

[The first identification of epidemic clone of enterotoxic Escherichia coli O∶6 serogroup highly associated with azithromycin resistance in Shanghai].

Objective: To investigate the molecular characterization of adult diarrhea cases caused by enterotoxic Escherichia coli (ETEC) and explore the practical model of epidemiology for laboratory technique and data needs based on the surveillance network. Methods: Epidemiological design and sampling targeted adult cases ETEC caused diarrhea in epidemic season. The enterotoxin type, serogroup, resistance, colonization factor and molecular type of ETEC were identified. Multiple dynamic phenotypic characteristics of ETEC were indicated by multidimensional and multivariable data. Results: From 2016 to 2018, 84 eligible ETEC strains were detected. The dominant serums/toxins were O∶6 (STh), O∶25 (LT), O∶159 (STh), O∶153 (STh). O∶6 (STh+CS21), which replaced O∶25 and O∶159 as the popular clones in 2018. Six cases of O∶153 (STh+CFA/I+CS8+PT34) in outbreak in 2017 were imported ones. The resistance rates of ETEC strains detected in adults to sulfasoxazole, naproxinic acid, ampicillin and azithromycin were more than 30%, multidrug resistance (MDR) reached 58.3%. Serum/toxin types suggested that attenuated strains were more likely to become MDR. Molecular typing confirmed that the genetic similarity of the dominant clone of O∶6 serogroup (PT20-24) was higher than O∶25 and O∶159. There was a high correlation between the minimal inhibitory concentration (MIC) of azithromycin and the resistant gene mphA (87.5%, 28/32). O∶6 (STh+CS21+mphA) resistant clone was first detected in 2016. Conclusion: A new epidemic clone in adult ETEC diarrhea cases in Shanghai was O∶6 (STh+CS21+mphA). For the first time the association between azithromycin resistance gene mphA and a serum group of ETEC was observed. Multidimensional and multivariate analysis techniques based on epidemiology can help reveal the potential transmission pattern of ETEC for the accurate surveillance and early warning of outbreaks.

Process risk model for Salmonella and ground chicken.

AIMS: To develop a process risk model (PRM) for evaluating the safety of individual lots of ground chicken (GC) contaminated with Salmonella (Salm). METHODS AND RESULTS: Data for prevalence, number and serotype of Salm were collected with 25 g samples of GC using a combination of methods (whole sample enrichment, quantitative polymerase chain reaction, cultural isolation and serotyping). These data were used to develop a predictive model for Salm contamination of GC as a function of serving size from 25 to 300 g. This model was combined with a model for thermal inactivation of Salm in GC and a dose-response model for Salm to develop a PRM in Excel that was simulated with NeuralTools and @Risk. Of 100, 25 g samples of GC examined, 19 tested positive for Salm. Three serotypes were isolated: Infantis (n = 13), Enteritidis (n = 5) and Typhimurium (n = 1). The number of Salm ranged from 0 to 2·56 log with a median of 0·93 log per 25 g of GC. The PRM predicted that Salm prevalence would increase (P < 0·05) from 19 to 57% to 82 to 93% as serving size increased from 25 to 100 g to 200 to 300 g. However, the total number of Salm in a 100-kg lot of GC and total severity of illness (TSI) were not affected (P> 0·05) by serving size. The PRM was also used to evaluate effects of serving size distribution, cooking, food consumption behaviour, consumer demographics and Salmonella virulence on TSI. CONCLUSIONS: How a lot of GC is partitioned and consumed does not affect TSI. Scenario analysis demonstrated that the PRM can integrate prevalence, number and serotype data for Salm with consumer handling, consumption and demographics data to identify safe and unsafe lots of GC for improved food safety and public health. SIGNIFICANCE AND IMPACT OF THE STUDY: Process-risk models like the one developed in this study represent a new, holistic approach to food safety that holds great promise for improving public health and reducing food recalls.

Molecular typing, antibiotic susceptibility, and biofilm production in nonencapsulated Streptococcus pneumoniae isolated from children in Japan.

The prevalence of nonencapsulated Streptococcus pneumoniae (NESp) has increased with the introduction of pneumococcal conjugate vaccines in children; however, the bacteriological characteristics of NESp have not been sufficiently clarified. In this study, NESp strains isolated from the nasopharyngeal carriage of children from four nursery schools in Japan were analyzed for molecular type, antibiotic susceptibility, and biofilm productivity. A total of 152 putative S. pneumoniae strains were identified by optochin-susceptibility analysis, of which 21 were not serotypeable by slide agglutination, quellung reaction, or multiplex PCR. Among these 21 strains, three were lytA-negative and, therefore, not S. pneumoniae. The remaining 18 strains were positive for lytA, ply, pspK, and bile solubility and were confirmed as NESp. Therefore, the isolation rate of NESp in the S. pneumoniae strains in this study was 12.0% (18/149). Molecular-typing analyses classified five strains as two existing sequence types (STs; ST7502 and ST7786), and 13 strains formed four novel STs. Horizontal spread was suspected, because strains with the same ST were often isolated from the same nursery school. The NESp isolates were generally susceptible to most antimicrobials, with the exception of macrolides; however, all isolates possessed more than one abnormal penicillin-binding protein gene. Furthermore, NESp strains were more effective than encapsulated counterparts at forming biofilms, which showed obvious differences in morphology. These data indicated that NESp strains should be continuously monitored as emerging respiratory pathogens.

Serotyping and antibiotic susceptibility patterns of Vibrio and Shigella isolates from diarrheal patients visiting a Tropical and Infectious Diseases Hospital in central Nepal.

BACKGROUND: Diarrheal diseases are the major infectious disease in developing countries like Nepal. Lack of proper sanitation and antimicrobial resistance gained by microbes have challenged to address diarrheal diseases in resource-limited countries. Early diagnosis of disease and proper antibiotic treatment can significantly reduce the disease burden. This study was designed to determine the recent antimicrobial susceptibility pattern of Vibrio cholerae and Shigella spp. to assure the proper antibiotic treatment. Stool specimens were processed following microbiological protocol and identified by biochemical and serological tests recommended by the Clinical Laboratory Standard Institute. RESULTS: Out of total 640 analyzed stool samples, 50 were culture positive, among them 29 were Shigella spp. (64.4%) and 21 were V. cholerae (46.6%). All V. cholerae strains belonged to the serogroup O1 and serovar Ogawa. Among the Shigella spp., Shigella flexneri 17 (59%) topped the list of serotype followed by Shigella sonnei 8 (28%), Shigella dysenteriae 3 (10%) and Shigella boydii 1 (3%) respectively. All the V. cholerae isolates (100%) were sensitive to cefotaxime while 71% were sensitive to tetracycline but 100 and 90.4% were resistance to co-trimoxazole and nalidixic acid respectively. Shigella isolates were mostly susceptible to cefotaxime (97%) while ciprofloxacin (48%) and ofloxacin (55%) were less effective drugs. CONCLUSIONS: These results on the prevalence of enteropathogens and their antibiotic resistance pattern may help to guide accurate choice of therapy in clinical setting. Hence, development of evidence based National Guidelines for the treatment of diarrhea is needed.

Non-typhoidal Salmonella in Calabria, Italy: a laboratory and patient-based survey.

INTRODUCTION: Although there has been a decrease in the number of cases of salmonellosis in the European Union, it still represents the primary cause of foodborne outbreaks. In Calabria region, data are lacking for the incidence of human non-typhoid salmonellosis as active surveillance has never been carried out. OBJECTIVE: To report the results of a laboratory and patient-based morbidity survey in Calabria to describe the incidence and distribution of Salmonella serovars isolated from humans, with a focus on antimicrobial resistance patterns. METHODS: Positive cultures from human samples were collected from every laboratory participating in the surveillance, with a minimum set of information about each isolate. A questionnaire was then administered to the patients by telephone interview to assess the potential risk exposures.Salmonella isolates underwent biochemical identification, molecular analysis by PCR and antimicrobial susceptibility testing by the disk-diffusion method. RESULTS: During a 2-year period, 105 strains of Salmonella spp were isolated from samples of patients with diarrhoea, with the highest isolation rate for children aged 1-5 years. The standardised rate was 2.7 cases per 1 00 000 population. The most common Salmonella isolates belonged to monophasic variant of S. Typhimurium (S. 4,[5],12:i:-) (33.3%), followed by S. Typhimurium (21.9%). 30.5% of the isolates were susceptible to all microbial agents tested and the most common pan-susceptible serotype was S. Napoli (100%). S. 4,[5],12:i:- was resistant to ampicillin, streptomycin, sulfonamides and tetracyclines in 42.9% cases, while resistance to quinolones was seen in 14.3% of the isolates. CONCLUSIONS: The results provide evidence that an active surveillance system effectively enhances Salmonella notifications. The high prevalence of antimicrobial resistance, including resistance to quinolones and multiresistance, enforces the need to strengthen strategies of surveillance and monitoring of antimicrobial use.

Increasing incidence of penicillin- and cefotaxime-resistant Streptococcus pneumoniae causing meningitis in India: Time for revision of treatment guidelines?

PURPOSE: Pneumococcal meningitis is a life-threatening infection, requiring prompt diagnosis and effective treatment. Penicillin resistance in pneumococcal infections is a concern. Here, we present the antibiotic susceptibility profile of pneumococcal meningeal isolates from January 2008 to August 2016 to elucidate treatment guidelines for pneumococcal meningitis. MATERIALS AND METHODS: Invasive pneumococcal isolates from all age groups, were included in this study. Minimum inhibitory concentrations for the isolates were identified by agar dilution technique and VITEK System 2. Serotyping of isolates was done by co-agglutination technique. RESULTS: Out of 830 invasive pneumococcal isolates, 167 (20.1%) isolates were from meningeal infections. Cumulative penicillin resistance in pneumococcal meningitis was 43.7% and cefotaxime non-susceptibility was 14.9%. Penicillin resistance amongst meningeal isolates in those younger than 5 years, 5-16 years of age and those aged 16 years and older was 59.7%, 50% and 27.3%, respectively, with non-susceptibility to cefotaxime in the same age groups being 18%, 22.2% and 10.4%. Penicillin resistance amongst pneumococcal meningeal isolates increased from 9.5% in 2008 to 42.8% in 2016, whereas cefotaxime non-susceptibility increased from 4.7% in 2008 to 28.5% in 2016. Serotypes 14, 19F, 6B, 6A, 23F, 9V and 5 were the most common serotypes causing meningitis, with the first five accounting for over 75% of resistant isolates. CONCLUSIONS: The present study reports increasing penicillin resistance and cefotaxime non-susceptibility to pneumococcal meningitis in our setting. This highlights the need for empiric therapy with third-generation cephalosporins and vancomycin for all patients with meningitis while awaiting results of culture and susceptibility testing.

Rapid Molecular Profiling of Defined Cell Types Using Viral TRAP.

Translational profiling methodologies enable the systematic characterization of cell types in complex tissues, such as the mammalian brain, where neuronal isolation is exceptionally difficult. Here, we report a versatile strategy for profiling CNS cell types in a spatiotemporally restricted fashion by engineering a Cre-dependent adeno-associated virus expressing an EGFP-tagged ribosomal protein (AAV-FLEX-EGFPL10a) to access translating mRNAs by translating ribosome affinity purification (TRAP). We demonstrate the utility of this AAV to target a variety of genetically and anatomically defined neural populations expressing Cre recombinase and illustrate the ability of this viral TRAP (vTRAP) approach to recapitulate the molecular profiles obtained by bacTRAP in corticothalamic neurons across multiple serotypes. Furthermore, spatially restricting adeno-associated virus (AAV) injections enabled the elucidation of regional differences in gene expression within this cell type. Altogether, these results establish the broad applicability of the vTRAP strategy for the molecular dissection of any CNS or peripheral cell type that can be engineered to express Cre.

Antibody Epitopes Identified in Critical Regions of Dengue Virus Nonstructural 1 Protein in Mouse Vaccination and Natural Human Infections.

Dengue is a global public health problem and is caused by four dengue virus (DENV) serotypes (DENV1-4). A major challenge in dengue vaccine development is that cross-reactive anti-DENV Abs can be protective or potentially increase disease via Ab-dependent enhancement. DENV nonstructural protein 1 (NS1) has long been considered a vaccine candidate as it avoids Ab-dependent enhancement. In this study, we evaluated survival to challenge in a lethal DENV vascular leak model in mice immunized with NS1 combined with aluminum and magnesium hydroxide, monophosphoryl lipid A + AddaVax, or Sigma adjuvant system+CpG DNA, compared with mice infected with a sublethal dose of DENV2 and mice immunized with OVA (negative control). We characterized Ab responses to DENV1, 2, and 3 NS1 using an Ag microarray tiled with 20-mer peptides overlapping by 15 aa and identified five regions of DENV NS1 with significant levels of Ab reactivity in the NS1 + monophosphoryl lipid A + AddaVax group. Additionally, we profiled the Ab responses to NS1 of humans naturally infected with DENV2 or DENV3 in serum samples from Nicaragua collected at acute, convalescent, and 12-mo timepoints. One region in the wing domain of NS1 was immunodominant in both mouse vaccination and human infection studies, and two regions were identified only in NS1-immunized mice; thus, vaccination can generate Abs to regions that are not targeted in natural infection and could provide additional protection against lethal DENV infection. Overall, we identified a small number of immunodominant regions, which were in functionally important locations on the DENV NS1 protein and are potential correlates of protection.

Serotype distribution and antimicrobial susceptibility of Streptococcus pneumoniae strains isolated in Japan after introduction of the routine immunization program.

Pneumococcal vaccines have reduced the incidences of Streptococcus pneumoniae infections among children and adults, but a relative increase in the prevalence of non-vaccine serotypes has been reported. To follow the changing epidemiology of pneumococcal diseases, capsular serotyping and antimicrobial susceptibility testing was performed on 534 pneumococcal isolates obtained from a hospital in Japan after routine immunization was launched, between October 2014 and May 2016. Serotype distributions and antimicrobial susceptibilities were evaluated among the total patient population, and were compared by age and sample groups and by serotype group, respectively. Serotypes targeted by the 13-valent pneumococcal conjugate vaccine (PCV13) were identified in 14.6%, 44.5%, and 40.2% of the samples from the <5, 5-64, and ≥65 year age groups, respectively. The 23-valent pneumococcal polysaccharide vaccine serotypes (PPSV23) were identified in 42.4%, 68.2%, and 63.1% of the samples, respectively; whereas non-PCV13 serotypes or non-PPSV serotypes (NVT) comprised 46.8% of all isolates. Among NVT, strain 35B was the most frequently isolated, followed by 15A, particularly in sputum samples collected from children <5 years old. Meanwhile, serotype 3, which is targeted by the PCV13 and PPSV23, was the most prevalent among patients aged ≥65 and 5-64 years. Antimicrobial susceptibility testing revealed that 88.9% and 81.0% of serotype 35B was non-susceptible to penicillin and meropenem, respectively, and 89.4% of 15A was non-susceptible to penicillin. Our data suggest rapid effects of pneumococcal vaccines and progression of serotype replacement. Besides invasive potential, the increased prevalence of non-vaccine serotypes with highly non-susceptible to penicillin was a concern. Continuous monitoring of pneumococcal serotypes and antimicrobial susceptibility is necessary for developing optimal preventive strategies.

Development of a Sensitive and Specific Polyclonal Antibody for Serological Detection of Clavibacter michiganensis subsp. sepedonicus.

The quarantine bacterium Clavibacter michiganensis subsp. sepedonicus (Cms) causes bacterial ring rot (BRR) in potato but is difficult to detect, hampering the diagnosis of this disease. ELISA immunoassays have not been widely used to detect Cms because commercially available anti-Cms antibodies detect mainly EPS-producing bacteria and can fail to detect strains that do not produce EPS. In the current study, we developed a new type of polyclonal antibody that specifically detects Clavibacter michiganensis subsp. sepedonicus bacteria irrespective of their EPS level. We first found that the presence of bacterial EPS precluded quantitative measurement of bacteria by currently available immunoenzymatic methods, but that washing Cms cells with acidic and basic buffers to remove EPS before analysis successfully standardized ELISA results. We used a mix of three strains of Cms with diverse EPS levels to generate antigen for production of antibodies recognizing Cms cells with and without an EPS layer (IgG-EPS and IgG-N-EPS, respectively). The resulting IgG-N-EPS recognized almost all Cms strains tested in this work regardless of their mucoidal level. The availability of this new antibody renders immunological diagnostics of Cms more sensitive and reliable, as our newly developed antibodies can be used in many type of immunoassays. This work represents an important step forward in efforts to diagnose and prevent the spread of BRR, and the methods and solutions developed in this work are covered by six Polish, one European and one US patents.

Antibacterial Activities of Ankaferd Hemostat (ABS) on Shiga Toxin-Producing Escherichia coli and Other Pathogens Significant in Foodborne Diseases.

OBJECTIVE: Ankaferd hemostat (Ankaferd Blood Stopper®, ABS)-induced pharmacological modulation of essential erythroid proteins can cause vital erythroid aggregation via acting on fibrinogen gamma. Topical endoscopic ABS application is effective in the controlling of gastrointestinal (GI) system hemorrhages and/or infected GI wounds. Escherichia coli O157:H7, the predominant serotype of enterohemorrhagic E. coli, is a cause of both outbreaks and sporadic cases of hemorrhagic colitis. The aim of this study is to examine the effects of ABS on 6 different Shiga toxigenic E. coli serotypes including O26, O103, O104, O111, O145, and O157 and on other pathogens significant in foodborne diseases, such as Salmonella Typhimurium, Campylobacter jejuni, and Listeria monocytogenes, were also assessed. MATERIALS AND METHODS: All strains were applied with different amounts of ABS and antimicrobial effect was screened. S. Typhimurium groups were screened for survival using the fluorescence in situ hybridization technique. RESULTS: The relative efficacy of ABS solutions to achieve significant logarithmic reduction in foodborne pathogens E. coli O157:H7 and non-O157 serogroups and other emerging foodborne pathogens is demonstrated in this study. ABS has antibacterial effects. CONCLUSION: Our present study indicated for the first time that ABS may act against E. coli O157:H7, which is a cause of thrombotic thrombocytopenic purpura, hemolytic-uremic syndrome, and hemorrhagic colitis. The interrelationships between colitis, infection, and hemostasis within the context of ABS application should be further investigated in future studies.

Invasive pneumococcal infections in children following transplantation in the pneumococcal conjugate vaccine era.

BACKGROUND: Pediatric recipients of hematopoietic stem cell and solid organ transplants are at increased risk of invasive pneumococcal infections (IPI). Data on IPI in this population are scarce. To our knowledge, this is the first study describing the epidemiology of IPI among pediatric transplant recipients in the pneumococcal conjugate vaccine (PCV) era. METHODS: We identified transplant recipients with IPI at 8 children's hospitals in the U.S. from our surveillance database (2000-2014). Pneumococcal isolates were collected prospectively. Serotyping and antibiotic susceptibility were performed in a central laboratory. Categorical variables were analyzed by Fisher's exact test and continuous variables with nonparametric tests. Indirect cohort study design was used to calculate vaccine effectiveness. RESULTS: We identified 65 episodes of IPI in transplant recipients. Recurrent IPI was observed in 10% of transplant recipients. The IPI crude incidence rate in solid organ transplant recipients was higher than in the general population. Most IPI episodes occurred >6 months after transplantation. Bacteremia and pneumonia were the most common presentations. Meningitis was unusual. No case fatalities were observed. Serotype 19A was the most common serotype (n=10), followed by 6C (n=7). In 2010-2014, 37% of IPI was caused by PCV13 serotypes. Four cases of vaccine breakthrough were identified. Most isolates were susceptible to penicillin and ceftriaxone. Pneumococcal conjugate and polysaccharide immunization rates were low. CONCLUSION: Pediatric transplant recipients remain at increased risk of IPI in the vaccine era. Most cases presented as a late post-transplant infection. The interval between transplantation and IPI may allow adequate time for pneumococcal immunization.

Antibiotic Resistance Patterns and Serotypes of Salmonella spp. Isolated at Jeollanam-do in Korea

OBJECTIVES: Few long-term studies have been conducted on the serotype and antibiotic resistance patterns of Salmonella speices (spp.) The aim of this study was to determine the serotypes and antibiotic resistance patterns of Salmonella spp. isolated at Jeollanam-do in Korea from 2004 to 2014. METHODS: A total of 276 Salmonella samples were evaluated. Serotyping was carried out according to the Kauffmann–White scheme. Antibiotic susceptibility was determined using the Vitek II system with an AST-N169 card. RESULTS: A total of 22 different serotypes were identified, and the major serotypes were Salmonella Enteritidis (116 strains, 42.0%) and Salmonella Typhimurium (60 strains, 21.7%). The highest resistance was observed in response to nalidixic acid (43.4%), followed by ampicillin (40.5%) and tetracycline (31.6%). Resistance to nalidixic acid was detected in 81.0% of S. Enteritidis. Multidrug resistance was detected in 43.3% of Salmonella spp. S. Enteritidis and S. Typhimurium presented the highest resistance (98.3%) and multidrug resistance rate (73.3%), respectively. The most highly observed antibiotic resistance pattern among Salmonella spp. in this study was ampicillin-chloramphenicol (14 strains, 5.7%). CONCLUSION: Overall, S. Enteritidis and S. Typhimurium showed higher antibiotic resistance than the other Salmonella serotypes tested in this study. Our study will provide useful information for investigating the sources of Salmonella infections, as well as selecting effective antibiotics for treatment.