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Glaesserella parasuis (GPS) is an important bacterial pathogen of swine. Serotype identification has presented a bottleneck in GPS research since it was first identified as the pathogen causing Glässer's disease in pigs in 1910. This paper presents a systematic review of the history of the development and application of gel immunodiffusion (GID), indirect hemagglutination assay (IHA), and polymerase chain reaction (PCR) typing methods for GPS, and the discovery of their shared antigenic basis. It provides a systematic theoretical overview of the immunology and principles underlying the three typing methods and offers new ideas for research into the prevention and control of Glässer's disease. In 1992, GPS was first classified into serotypes 1–15 using GID based on GPS heat-stable antigens, but about 25% of the strains were found to be non-typeable, and the composition of their antigens for serotyping was unclear. In 2003, the IHA method was established based on saline-extracted antigens of GPS, whose sensitivity and typing rate were higher than for GID, although about 15% of strains were still found to be non-typeable. The results of IHA and GID typing are roughly consistent, since they share the same GPS surface polysaccharide serotyping antigens, although whether these are capsular polysaccharides, lipopolysaccharides, or other polysaccharides, remains to be determined. In 2013, the Capsular polysaccharide (CPS) synthetic gene clusters from GPS serotypes 1–15 were successfully analyzed, confirming that CPS is essential for the formation of antigens for serotyping. In 2015, primers were designed based on the specific target genes of GPS capsules to establish a PCR typing method (H-PCR) for GPS, which, however, could not identify serotypes 5 and 12. In 2017, a new PCR typing method (J-PCR) was established based on the specific target genes of GPS capsules, which could identify serotypes 5 and 12. A combination of the two PCR typing methods enables the typing of almost all GPS strains, and the consistency with GID and IHA was verified using molecular biological methods. The antigenic basis of the three typing methods was shown to involve the GPS capsule. PCR typing methods are characterized by simple operation, fast speed, and low cost, and can successfully solve many problems in GID and IHA serotyping, and so have become widely adopted. [ABSTRACT FROM AUTHOR]

This study investigated the presence of Pasteurella multocida in Egyptian dairy farm calves with respiratory symptoms. A total of 150 samples were collected aseptically from 133 nasal swabs and 17 pneumonic lung tissues of calves on private farms across different governorates. Samples were tested using three methods: standard bacteriological examination, biochemical identification, and Molecular characterization for P. multocida. The P. multocida isolates were subjected to multiplex PCR for capsule biosynthesis and 16S rRNA gene. Capsular typing revealed that 60 out of 150 samples (40%) were positive for P. multocida. All isolates confirmed as P. multocida through PCR were belonged to serogroup A. Antibiotic sensitivity testing of these isolates identified enrofloxacin as the most effective drug. The most detectable resistance gene was sul II-in examined isolates with a 100% detection rate, followed by BlaTEM and tetA genes (62.5%). Histopathological examination revealed extensive damage across various organs, such as the lungs of calves, including fibrinous exudate inside alveoli and purulent fluid in other alveoli, besides thrombosis of blood vessels in P. multocida-infected animals. [ABSTRACT FROM AUTHOR]

Streptococcus agalactiae, or group B streptococcus (GBS), is a Gram-positive pathogen with an extended track record of colonization in the gastrointestinal and genitourinary tracts. GBS can induce disease in individuals across all age demographics, yet it predominantly triggers infections in neonates and the elderly. Identification of the serotype is vital for effective management of the disease as it provides critical information for clinicians on the cause of the disease. In this study, we evaluated the rapid, simple, and easy-to-adopt multiplex real-time PCR technique, NovaplexTM (NovaPCR). A total of 131 clinical isolates of different serotypes were tested using NovaPCR. Observations revealed that 129 isolates showed the same observations as LA and conventional mPCR. NovaPCR accurately identified serotypes IV and V, which were first classified as serotype Ia in the LA test and mPCR, and the difference between the traditional (LA test and mPCR) and NovaPCR methods is only 1.52%. Accurate serotype identification is helpful for monitoring the epidemics and achieving optimal clinical outcomes, and NovaPCR showed a reliable, fast, easy-to-interpret, and cost-efficient performance in GBS serotyping. [ABSTRACT FROM AUTHOR]

The present study describes the genetic characterization of L. monocytogenes strains found in the Republic of Kosovo's food chain. From 2016 to 2022, 995 samples were collected. Overall, 648 samples were from ready-to-eat (RTE) food products, 281 from food products consumed cooked (FPCC), 60 from raw materials, and 6 from environmental samples. Overall, 11.76% (117 out of 995) of the samples were contaminated by L. monocytogenes, comprising 6.33% (41 out of 648) from RTE products, 14.95% (42 out of 281) from FPCC, 55.00% (33 out of 60) from raw materials, and 16.66% (1 out of 6) from environmental samples. All isolates were subjected to molecular serotyping and clonal complex (CC) identification by using real-time PCR, as well as multilocus sequence typing. All isolates were grouped into four molecular serotypes, IIa (34.19%), IIb (3.48%), IIc (32.48%), and IVb (29.91%), as well as Lineage I (33.33%) and Lineage II (66.66%). In total, 14 CCs were identified from 41 RTE isolates; however, CC29 (7), CC2 (6), and CC6 (6) were the most dominant. By contrast, CC9 was by far the most represented CC in both FPCC (21) and RM (14). Moreover, 30 isolates expressed CC1, CC2, CC4, or CC6, which are particularly associated with severe human infections. [ABSTRACT FROM AUTHOR]

Due to their immobile nature and filter-feeding habits, which enable biological particles to accumulate in their tissues, mussels are recognized as vectors of foodborne diseases. Consuming these shellfish uncooked or partly processed might result in food poisoning because of accumulated bacteria originating from the culture environment and unhygienic handling. The present study aimed to assess the presence of Salmonella and Escherichia coli as well as to biochemically and serotypically confirm Salmonella spp. in mussels (Mytilus galloprovincialis) taken from two locations in Morocco: Zenata and Mansouria. A total of 90 mussel samples were collected from October 2022 to August 2023. Two methods were employed in this study: AFNOR NF EN ISO 6579-1 (2017) for the detection of Salmonella spp., and the most probable number method (MPN) using Norm ISO/TS 16649-3 (2017) for E. coli. The number of E. coli varied between 0.2/100 g and 1.7 x 103/100 g of mussels. The percentage of Salmonella spp. detected in mussel samples was 4,4%. Further analysis revealed the identification of two distinct Salmonella serotypes, namely S. kentucky (1 isolate) and S. Typhimurium (3 isolates). This research highlights the potential risks to public health due to the presence of pathogenic bacteria in mussels from two regions of Morocco where shellfish farming and coastal tourism are significant contributors to the local economy. [ABSTRACT FROM AUTHOR]

Since 1998, notifiable bluetongue virus (BTV) serotypes 1–4, 6, 8, 9, 11, and 16 have been reported in Europe. In August 2006, a bluetongue (BT) outbreak caused by BTV serotype 8 began in northwestern Europe. The Netherlands was declared BT-free in February 2012, and annual monitoring continued. On September 3, 2023, typical BT clinical manifestations in sheep were notified to the Netherlands Food and Product Safety Consumer Authority. On September 6, we confirmed BTV infection through laboratory diagnosis; notifications of clinical signs in cattle were also reported. We determined the virus was serotype 3 by whole-genome sequencing. Retrospective analysis did not reveal BTV circulation earlier than September. The virus source and introduction route into the Netherlands remains unknown. Continuous monitoring and molecular diagnostic testing of livestock will be needed to determine virus spread, and new prevention strategies will be required to prevent BTV circulation within the Netherlands and Europe.

ObjectiveTo provide scientific basis for the prevention and control， early warning， tracing and clinical treatment of diarrheal diseases caused by Salmonella， the serotype distribution， drug resistance and molecular typing characteristics of Salmonella isolates from infectious diarrhea patients in Huzhou City were analyzed.MethodsA total of 126 isolates of Salmonella were collected from 6 sentinel hospitals in Huzhou City from 2021 to 2022. The isolates were characterized by serovar determination， antimicrobial susceptibility tests， and pulsed-field gel electrophoresis （PFGE） typing.ResultsThe 126 strains of Salmonella could be divided into 28 serotypes. The dominant serotypes were Salmonella typhimurium monophasic variant （32.5%， 41/126）， Salmonella enteritidis （24.6%， 31/126） and Salmonella typhimurium （15.1%， 19/126）. Salmonella typhimurium had different resistance characteristics to 15 antibiotics. The resistance rate of Salmonella typhimurium monophasic variant to tetracycline was 73.3% （44/60）， ampicillin and sulfamethoxazole were all 56.7% （34/60）. The resistance rate of Salmonella typhimurium to tetracycline and sulfamethoxazole was higher with 68.4% （13/19） and 52.6% （10/19）， respectively. The highest resistance rate of Salmonella enteritidis to ampicillin/sulbactam was 54.8% （17/31） and that of other Salmonella to tetracycline was the highest wih 51.4%（18/35）. The overall multidrug resistance rate was 62.7% （79/126）， showing 20 multidrug resistance spectra. The PFGE fingerprints of Salmonella typhimurium and Salmonella enteritidis were obtained 46 and 15 kinds types after digested by Xba Ⅰ enzyme.ConclusionThe predominant serotypes of Salmonella infection in Huzhou City are Salmonella typhimurium and Salmonella enteritidis. The multi-drug resistance of Salmonella bacteria is serious. PFGE zone types are both diverse and dominant.

Objective: To describe the profile of Streptococcus pneumoniae, identify research gaps, and provide in‐depth insights into various aspects related to the pathogen. Methods: Google Scholar, PubMed, and ScienceDirect were searched for all studies on the pneumococcus in Ghana that reported on specimen collected, population and sample size, carriage prevalence, incidence of pneumococcal diseases, age of the study population, types of test performed, serotypes identified, antimicrobial susceptibilities, or molecular analysis on the pneumococci for data extraction. Results: Overall, a total of 7954 results were obtained from the three‐database search, and of this, 24 articles were selected after screening. A total of 924 isolates were accounted for by serotyping/serogrouping. The prevalence of pneumococcal carriage in Ghana ranges from 11.0% to 51.4% in the population depending on the age (≤ 24–80 years), sickle cell disease (SCD), human immunodeficiency virus (HIV) status, or health of the study population, and penicillin (Pen)‐nonsusceptible isolates ranged from 17% to 63%. The prevalence of pneumococci found as the etiologic agent of diseases among Ghanaians ranges from 3.4% for otitis media to 77.7% for meningitis. Overall, the 13‐valent pneumococcal conjugate vaccine (PCV) (PCV‐13) carriage serotypes accounted for 28.4% of the reported pneumococcal isolates. PCV‐13 invasive serotypes accounted for 22.4% of the reported isolates. The non‐PCV‐13 carriage serotypes accounted for most (43.9%) of the reported isolates. In the pre‐PCV‐13 era, the nontypeable (NT) (5.5%) and other nonvaccine types (NVTs) (6.4%) were reported as being predominant. The non‐PCV‐13 serotypes accounted for 4.4% of the reported isolates in invasive pneumococcal disease (IPD) cases. Multidrug resistance (MDR) ranged from 7.8% to 100%. Conclusion: Predicting the invasiveness of pneumococci using molecular typing is the way to go in the future as this will provide answers to the extent to which capsular switching is contributing to the pneumococcal disease burden in Ghana almost a decade after introducing PCV‐13. Continuous monitoring of antibiotic resistance patterns at both phenotypic and genotypic levels, along with serotyping and molecular typing, should be a standard practice in the surveillance of pneumococcal disease burden in Ghana. [ABSTRACT FROM AUTHOR]

Simple Summary: Porcine pleuropneumonia, caused by Actinobacillus pleuropneumoniae, is one of the main causes of massive losses in the pig industry, with a high economic impact. This study aimed to identify A. pleuropneumoniae serotypes isolated from pneumonic lesions in dead pigs, naturally infected, in some Piedmontese farms and to describe the macroscopic and microscopic lesions. The identification of circulating serotypes in swine farming is crucial to adopting optimal strategies to control the diseases, such as vaccination plans and biosecurity measures. Porcine pleuropneumonia (PPP) is one of the main causes leading to massive losses in the pig industry, with high economic impacts. Among different etiological agents, Actinobacillus pleuropneumoniae (APP) is responsible for severe fibrinous-necrotizing pleuropneumonia. A total of 19 different APP serotypes are currently recognized. This study aimed to identify APP serotypes isolated from pneumonic lesions in naturally infected and dead pigs in the Piedmont Region and to describe lesions. A total of 107 dead pigs with a suspected PPP diagnosis were included in this study. Lungs were evaluated using gross-pathology scoring systems, histopathology, and APP isolation and serotypes identification by multiplex PCR were conducted. Gross lung lesions were mainly represented by fibrinous pneumonia and pleuropneumonia. APP was isolated in 20/107 (18.7%) samples. PCR indicated APP DNA presence in 53/107 (49.5%) of lung samples. The most observed serotypes were serotype 2 in 24/53 (45.3%) and serotype 6 in 13/53 (24.5%) samples. Moreover, multiplex PCR results suggested a coinfection of different serotypes in five samples. This study emphasizes the importance of an integrated approach, utilizing various techniques, such as gross- and histopathology, and bacteriological culture and PCR, to enhance the diagnosis of APP infections. [ABSTRACT FROM AUTHOR]

There is scant information on Salmonella in indigenous poultry in Nigeria. We investigated the occurrence and characterized Salmonella serovars in indigenous poultry and their drinking water sources to enhance the monitoring of the infection in poultry and to promote public health. We collected 1208 samples, poultry droppings (n = 1108), and water (n = 100) across 15 markets in North Central Nigeria. Salmonella spp. were isolated following World Organisation for Animal Health guidelines. Salmonella spp., confirmed through invA gene detection by a polymerase chain reaction assay, were 6.8% (75/1108) droppings and 3% (3/100) water. Susceptibility testing against 13 antimicrobials showed 60.3% (47/78) susceptibility to all the antimicrobials tested while 14.1% (11/78) were multidrug resistant. Serotyping and whole-genome sequencing were carried out on 44 of the isolates, and 23 different serovars were identified. Genomes of serovars Luedinghausen, Laredo, Widemarsh, and Lansing are being documented in Africa for the first time. Twenty (20) antimicrobial resistance (AMR) gene markers encoding for resistance to aminoglycosides, tetracyclines, sulphonamides, quinolones, trimethoprim, penicillins and phenicols were found. Phylogenetic cluster analysis showed close relatedness among isolates from different sources. This study shows both low Salmonella prevalence and AMR, but since uncommon serovars are circulating, continuous monitoring is recommended so as to ensure food safety and poultry health. [ABSTRACT FROM AUTHOR]

• Carriage of vaccine serotypes declined following PCV10 introduction. • Multiple-serotype carriage was reduced following PCV10 introduction. • Pneumococcal carriage density among children was lower following PCV10 introduction. Carriage studies are an efficient means for assessing pneumococcal conjugate vaccine effect in settings where pneumococcal disease surveillance programmes are not well established. In this study the effect of 10-valent pneumococcal conjugate vaccine (PCV10) introduction on pneumococcal carriage and density among Nepalese children using a bacterial microarray and qPCR was examined. PCV10 was introduced into the Nepalese infant immunisation schedule in August 2015. Nasopharyngeal swabs were collected from healthy Nepalese children in Kathmandu between April 2014 and December 2021. Samples were plated on blood agar, incubated overnight, and DNA extracted from plate sweeps. Pneumococcal serotyping was done using the Senti-SPv1.5 microarray (BUGS Bioscience, UK). DNA was extracted from swab media and qPCR performed for pneumococcal autolysin (lytA). A significant decline in prevalence of PCV10 serotypes was observed when comparing pre-PCV10 with post-PCV10 collection periods (36.5 %, 454/1244 vs 10.3 %, 243/2353, p < 0.0001). Multiple-serotype carriage was also observed to significantly decline when comparing pre-PCV10 with post-PCV10 periods (31.4 %, 390/1244 vs 22.2 %, 522/2353, p < 0.0001). Additionally, a significant decline in median pneumococcal density was observed when comparing pre-PCV10 with post-PCV10 periods (3.3 vs 3.25 log10 GE/ml, p = 0.0196). PCV10 introduction was associated with reduced, prevalence of all PCV10 serotypes, multiple serotype carriage, and pneumococcal carriage density. [ABSTRACT FROM AUTHOR]

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Anamnestic 13-valent pneumococcal conjugate vaccine immunization did not affect the relapse risk in pediatric idiopathic nephrotic syndrome. Pneumococcal serotype (PS)-specific antibody titers increased significantly in all groups. Children receiving immunomodulatory treatments (IMTs) displayed significantly lower levels of PS-specific antibodies for 3/8 serotypes tested. PS-specific B-cell counts significantly increased only in healthy controls and patients receiving corticosteroids. [ABSTRACT FROM AUTHOR]

Small ruminant lentiviruses (SRLV) cause multisystemic, degenerative and chronic disease in sheep and goats. There are five genotypes (A, B, C, D and E), of which A and B are the most widespread. The purpose of this study was to evaluate the serotyping efficiency of the Eradikit SRLV Genotyping ELISA and the molecular typing efficiency of a newly developed nested real-time PCR targeting the long terminal repeat–gag (LTR-gag) region using samples from animals infected with subtypes of SRLV known to circulate in Poland. A total of 97 sera samples taken from 34 sheep and 63 goats were immunoassayed, and 86 DNA samples from 31 sheep and 55 goats were tested with the PCR. All ruminants were infected with known SRLV strains of the A1, A5, A12, A13, A16, A17, A18, A23, A24, A27, B1 and B2 subtypes. A total of 69 (80.2%, 95% confidence interval 71.6%–88.8%) out of 86 tested samples gave positive results in the PCR. In 17 out of the 86 (19.8%) samples, no proviral DNA of SRLV was detected. The differentiation between MVV (genotype A) and CAEV (genotype B) by PCR matched the predating phylogenetic analysis invariably. No cross-reactivity was observed. On the other hand, the proportion of samples genotyped the same by the older phylogenetic analysis and the Eradikit SRLV Genotyping ELISA was 42.3%. The test was unable to classify 40.2% of samples, and 17.5% of sera were incorrectly classified. Our results showed that the Eradikit SRLV genotyping kit is not a reliable method for predicting SRLV genotype, while the nested real-time PCR based on the LTR-gag region did prove to be, at least for genotypes A and B. [ABSTRACT FROM AUTHOR]

Objective To analyze the serotypes, drug resistance, and molecular epidemiological characteristics of non-typhoidal Salmonella in Chaozhou City, Guangdong Province in 2022, providing pathogenic support for the effective prevention and control of foodborne diseases. Methods In 2022, human-derived non-typhoidal Salmonella strains and food-derived strains were isolated through sentinel surveillance and food risk monitoring in Chaozhou City, Guangdong Province. The strains were subjected to serotyping, drug susceptibility testing, and whole-genome sequencing analysis. Results A total of 1 257 fecal specimens were collected for diarrhea surveillance in Chaozhou City in 2022, and 58 strains of non-typhoidal Salmonella were detected, with an isolation rate of 4.61%. Among 116 foodstuffs, 2 strains of non-typhoidal Salmonella were detected, with an isolation rate of 1.72%. Early childhood was the main age group for infection (14/144, 9.72%). The highest detection rate was in June (12/123, 9.76%). The 58 human-derived non-typhoidal Salmonella strains were classified into 12 serotypes, with Salmonella typhimurium being the most prevalent (44/58, 75.86%). In terms of drug resistance, 55 strains showed resistance, with a resistance rate of 91.67%, among which ampicillin, tetracycline, chloramphenicol, and streptomycin had the highest resistance rates, 76.67%, 70.00%, 67.67%, and 60.00%, respectively. Forty-nine strains were multi-resistant, accounting for 81.67%. From the molecular characteristics standpoint, multilocus sequence typing (MLST) divided the strains into 14 sequence typing (ST), with Salmonella typhimurium ST34 being the dominant type. Whole-genome single nucleotide polymorphism (wg-SNP) evolutionary analysis revealed high genetic heterogeneity among the studied strains, with Salmonella typhimurium clustering together and showing close evolutionary relationships to clinical samples of Salmonella enteritidis. Conclusions In 2022, the non-typhoidal Salmonella strains from Chaozhou showed a diversity of serotypes, with Salmonella typhimurium being predominant and exhibiting severe multidrug resistance. Molecular typing revealed diverse evolutionary characteristics. [ABSTRACT FROM AUTHOR]

BACKGROUND: Nontypeable Haemophilus influenzae (NTHi) is the most common cause of invasive H. influenzae disease in the United States (US). We evaluated the epidemiology of invasive NTHi disease in the US, including among pregnant women, infants, and people with human immunodeficiency virus (PWH). METHODS: We used data from population- and laboratory-based surveillance for invasive H. influenzae disease conducted in 10 sites to estimate national incidence of NTHi, and to describe epidemiology in women of childbearing age, infants aged ≤30 days (neonates), and PWH living in the surveillance catchment areas. H. influenzae isolates were sent to the Centers for Disease Control and Prevention for species confirmation, serotyping, and whole genome sequencing of select isolates. RESULTS: During 2008-⁠2019, average annual NTHi incidence in the US was 1.3/100 000 population overall, 5.8/100 000 among children aged

ABSTRACT Klebsiella pneumoniae has emerged as a global health threat due to its role in the spread of antimicrobial resistance and because it is a frequent cause of hospital-acquired infections and neonatal sepsis. Capsular and lipopolysaccharide (LPS) O-antigen polysaccharide surface antigens are major immunogens that are useful for strain classification and are candidates for vaccine development. We have developed real-time PCR reagents for molecular serotyping, subtyping, and quantitation of the most prevalent LPS O-antigen types (i.e., O1, O2, O3, and O5) of Klebsiella pneumoniae. We describe two applications for this O-typing assay: for screening culture isolates and for direct typing of Klebsiella pneumoniae present in stool samples. We find 100% concordance between the results of the O-typing assay and whole-genome sequencing of 81 culture isolates, and >90% agreement in O-typing performed directly on specimens of human stool, with disagreement arising primarily from a lack of sensitivity of the culture-based comparator method. Additionally, we find evidence for mixed O-type populations at varying levels of abundance in direct tests of stool from a hospitalized patient population. Taken together, these results demonstrate that this novel O-typing assay can be a useful tool for K. pneumoniae epidemiologic and vaccine studies.IMPORTANCEKlebsiella pneumoniae is an important opportunistic pathogen. The gastrointestinal (GI) tract is the primary reservoir of K. pneumoniae in humans, and GI carriage is believed to be a prerequisite for invasive infection. Knowledge about the dynamics and duration of GI carriage has been hampered by the lack of tools suitable for detection and strain discrimination. Real-time PCR is particularly suited to the higher-throughput workflows used in population-based studies, which are needed to improve our understanding of carriage dynamics and the factors influencing K. pneumoniae colonization.

ObjectiveTo provide scientific basis for rational clinical use and traceability of outbreaks of foodborne Salmonellosis， the drug resistance and molecular typing characteristics of foodborne Salmonella in Ganzhou City was understood， and a database of drug resistance and molecular fingerprinting of foodborne Salmonella in Ganzhou City was established.MethodsSerotyping， drug susceptibility testing， whole genome sequencing （WGS） and pulsed field gel electrophoresis （PFGE） were performed on 136 strains of Salmonella isolated from the active surveillance of foodborne diseases in Ganzhou City from 2020 to 2022. And drug resistance gene annotation and graphical clustering analysis were performed.ResultsThe highest rate of foodborne Salmonella in Ganzhou City was found for STR resistance （83.09%）， followed by TET （78.68%） and AMP （76.47%）. Multi-drug resistant strains accounted for 76.47%， with a wide range of resistance spectrum types， and the main prevalent resistance spectrum type was AMP-TET-CHL-STR-SXT. WGS predicted a total of 61 resistance genes in 7 categories， with the highest rate of aminoglycoside resistance genes （99.19%） and the lowest rate of macrolides （8.87%）. 136 strains of Salmonella with S. typhimurium variant and S. typhimurium as the dominant serotype were classified into 98 band types by PFGE molecular typing.ConclusionFoodborne Salmonella in Ganzhou City has a serious drug resistance status， with a high rate of drug resistance gene carriage and diverse genotypes. The PFGE molecular type is polymorphic. The dominant serotype may cause outbreaks and epidemics， and surveillance and research should be strengthened.

Background and Aim: Non-Typhoidal Salmonellae (NTS) is emerging as a major health problem globally. They cause foodborne infections, especially gastroenteritis as well as extraintestinal manifestations. As this is a zoonotic disease, certain animal sources such as poultry, egg, meat and dairy products are implicated. Fruits and vegetables too contaminated by manure of infected animal excreta are possible vehicles of transmission of this disease. The present report analyzes the spectrum of NTS with seroprofiling, isolated in our institute for the last 12 years. Materials and Methods: Fresh fecal samples from patients with acute gastroenteritis and blood samples from cases of bacteremia were processed according to the standard protocol. Nonlactose fermenting colonies resembling Salmonella were subjected to biochemical characterization and antimicrobial susceptibility testing followed by serotyping. Results: One hundred fifty-two NTS strains were isolated from feces and 13 from blood. Serotyping of these strains revealed that the most common NTS species was Salmonella Typhimurium (44.8%), followed by Salmonella Oslo (15.2%) and Salmonella Weltevreden (11.5%). Many rare strains were also encountered in our study. Conclusion: As many of the NTS species are biochemically similar, serotyping of NTS should be made mandatory to know the burden of the disease and to detect the emergence of new serovars. Emergence of multidrug resistance in NTS strains is a major concern nowadays. It is very essential to adopt strict quality measures in poultry, fish, dairy, and meat processing units as they are widely distributed in animals. In addition, strategies should be formulated by health education and public awareness programs to sensitize the population regarding this menace.

Salmonella spp. remains a significant concern in poultry flocks due to its potential impact on both animal health and public health. In this study, a comprehensive approach to isolate, serotype, and identify Salmonella strains from poultry flocks was employed. A total of 350 samples were collected, including internal organs liver, lung and cecum, from poultry flocks. Salmonella isolates were isolated and characterized according to established protocols. Conventional serotyping was performed based on the Kauffman–White scheme using Salmonella antisera, and the results were classified into various serogroups. To enhance the efficiency of serotyping, a multiplex PCR assay targeting both O and H antigens was developed. DNA extraction from collected samples followed the boiling method the multiplex PCR assay used specific primers for various serogroups. Out of the 350 samples tested, approximately 14.2% were positive for Salmonella. A total of 50 Salmonella isolates were serotyped, and the strains were categorized into several serogroups including B, D1, E1, C1, and C3. The multiplex PCR assay successfully identified O and H antigens, revealing a prevalence of serogroup B (46%) and D1 (32%) strains. Interestingly, none of the Salmonella strains exhibited the Vi antigen. This investigation showcased the precision and effectiveness of the multiplex PCR technique in identifying Salmonella strains. Notably, the multiplex PCR yielded outcomes surpassing those of traditional serotyping using the Kaufmann White scheme, all within a notably reduced time frame. In conclusion, the study highlights and presents a rapid and reliable method for serotyping and identifying Salmonella strains using multiplex PCR. This approach has the potential to expedite the control and prevention of Salmonella transmission, contributing to improved poultry health and reduced public health risks associated with Salmonella infections. The rapidity and accuracy of the multiplex PCR method make it particularly suitable for international efforts to monitor and manage Salmonella outbreaks effectively

The clinical consequences of toxoplasmosis are greatly dependent on the Toxoplasma gondii strain causing the infection. To better understand its epidemiology and design appropriate control strategies, it is important to determine the strain present in infected animals. Serotyping methods are based on the detection of antibodies that react against segments of antigenic proteins presenting strain-specific polymorphic variations, offering a costeffective, sensitive, and non-invasive alternative to genotyping techniques. Herein, we evaluated the applicability of a panel of peptides previously characterized in mice and humans to serotype sheep and pigs. To this end, we used 51 serum samples from experimentally infected ewes (32 type II and 19 type III), 20 sheep samples from naturally infected sheep where the causative strain was genotyped (18 type II and 2 type III), and 40 serum samples from experimentally infected pigs (22 type II and 18 type III). Our ELISA test results showed that a combination of GRA peptide homologous pairs can discriminate infections caused by type II and III strains of T. gondii in sheep and pigs. Namely, the GRA3-I/III-43 vs. GRA3-II-43, GRA6-I/III-213 vs. GRA6-II-214 and GRA6-III- 44 vs. GRA6-II-44 ratios showed a statistically significant predominance of the respective strain-type peptide in sheep, while in pigs, in addition to these three peptide pairs, GRA7-II-224 vs. GRA7-III-224 also showed promising results. Notably, the GRA6-44 pair, which was previously deemed inefficient in mice and humans, showed a high prediction capacity, especially in sheep. By contrast, GRA5-38 peptides failed to correctly predict the strain type in most sheep and pig samples, underpinning the notion that individual standardization is needed for each animal species. Finally, we recommend analyzing for each animal at least 2 samples taken at different time points to confirm the obtained results. [ABSTRACT FROM AUTHOR]

Background and Aim: Influenza A subtypes H3N2 and H1N1 are currently circulating among humans and are the main cause of seasonal epidemics in the temperate regions. The specific clinical features of influenza H1N1 and H3N2 infections have not been well evaluated in Iranian patients. Materials and Methods: An observational study was conducted on influenza-confirmed patients in Imam Khomeini Hospital Complex, Tehran, Iran from October 2022 to March 2023. The specimens tested positive for influenza, using RT-PCR, were transported to the National Influenza Center (NIC) Laboratory for the typing identification. Different data, including demographics, comorbidities, length of hospitalization, outcomes, paraclinical features, and recent influenza vaccination, were collected. Results: A number of 165 hospitalized patients with influenza infection were included: 148 (89.70%) patients with H3N2 and 17 (10.30%) with H1N1 infections. Among the patients 55.20% were male. The median age was 49 years. The patients were symptomatic for an average of 8 days. The most common symptoms were dyspnea (60.6%), fever (58.8%), and productive cough (37.6%). The median oxygen saturation was significantly lower in the H3N2 group (91%–81.0, 96.0) than in the H1N1 group (95%–90.0, 97.5) (P=0.029). The ground glass opacity (GGO) was the most common (69.1%) pulmonary radiological pattern. Only one patient had received the influenza vaccine in the recent year.. The mortality rate (19 patients: 11.55%) was associated with the cancer and liver diseases. Conclusion: Influenza A (H3N2 and H1N1) can be considered the outbreak cause in Iran in 2022–2023. The vaccination rate was very low and the mortality rate was high. Influenza vaccination is recommended to prevent death, especially in the patients with cancer. [ABSTRACT FROM AUTHOR]

Salmonella enterica, a bacterium causing foodborne illnesses like salmonellosis, is prevalent in Europe and globally. It is found in food, water, and soil, leading to symptoms like diarrhea and fever. Annually, it results in about 95 million cases worldwide, with increasing antibiotic resistance posing a public health challenge. Therefore, it is necessary to detect and serotype Salmonella for several reasons. The identification of the serovars of Salmonella enterica isolates is crucial to detect and trace outbreaks and to implement effective control measures. Our work presents a protein-based microarray for the rapid and accurate determination of Salmonella serovars. The microarray carries a set of antibodies that can detect different Salmonella O- and H-antigens, allowing for the identification of multiple serovars, including Typhimurium and Enteritidis, in a single miniaturized assay. The system is fast, economical, accurate, and requires only small sample volumes. Also, it is not required to maintain an extensive collection of sera for the serotyping of Salmonella enterica serovars and can be easily expanded and adapted to new serovars and sera. The scientific state of the art in Salmonella serotyping involves the comparison of traditional, molecular, and in silico methods, with a focus on economy, multiplexing, accuracy, rapidity, and adaptability to new serovars and sera. The development of protein-based microarrays, such as the one presented in our work, contributes to the ongoing advancements in this field. [ABSTRACT FROM AUTHOR]

Listeria monocytogenes (L. monocytogenes) poses a significant threat to food safety due to its ability to cause severe human illness and its resistance to various antibiotics and environmental conditions. This study investigated the prevalence, serotype distribution, virulence gene profiles, and antimicrobial resistance patterns of L. monocytogenes in ready-to-eat (RTE) food products from Romania. A total of 8151 samples were analyzed, including various processed dairy, bovine, poultry, pork, and fish products. Bacterial isolation was conducted using the classical standard method, followed by confirmation through biochemical and molecular testing. Among the isolated strains, serotypes 1/2a, 1/2b, and 1/2c were identified, with a prevalence of 75% for serotype 1/2a. Additionally, virulence genes specific to listeriolysin O (hlyA) and regulatory factor A (prfA) were detected in all isolates. Antimicrobial susceptibility testing revealed varying resistance patterns among the L. monocytogenes strains. Trimethoprim-sulfamethoxazole and oxacillin showed the highest prevalence of resistance at 26.92% and 23.07%, respectively. However, all strains remained susceptible to ciprofloxacin, levofloxacin, and moxifloxacin. Notably, 23.07% of the isolates exhibited multidrug resistance, with the most common pattern being resistance to oxacillin, penicillin, and tetracycline. Analysis of antimicrobial resistance genes identified tetracycline resistance genes, particularly tet(C), tet(M), and tet(K), in a significant proportion of isolates. The presence of ampC and dfrD genes was also notable, indicating potential mechanisms of resistance. These results emphasize the necessity for ongoing surveillance of L. monocytogenes in RTE foods and emphasize the importance of thorough monitoring of antimicrobial resistance to guide public health strategies within the European Union. [ABSTRACT FROM AUTHOR]

African horse sickness is a severe and often fatal disease affecting all species of equids. The aetiological agent, African horse sickness virus (AHSV), can be differentiated into nine serotypes. The identification of AHSV serotypes is vital for disease management, as this can influence vaccine selection and help trace disease incursion routes. In this study, we report the development and optimisation of a novel, molecular-based assay that utilises multiplex PCR and microsphere-based technology to expedite detection and differentiation of multiple AHSV serotypes in one assay. We demonstrated the ability of this assay to identify all nine AHSV serotypes, with detection limits ranging from 1 to 277 genome copies/µL depending on the AHSV serotype. An evaluation of diagnostic sensitivity and specificity revealed a sensitivity of 88% and specificity of 100%. This method can serotype up to 42 samples per run and can be completed in approximately 4–6 h. It provides a powerful tool to enhance the rapidity and efficiency of AHSV serotype detection, thereby facilitating the generation of epidemiological data that can help understand and control the incidence of AHSV worldwide. [ABSTRACT FROM AUTHOR]

Simple Summary: Small ruminant lentiviruses are a group of viruses characterized by a high genetic and antigenic variability. In the absence of control measures, they are characterized by high prevalence in goat and sheep farms. In Greece, a country characterized by one of the highest small ruminant populations in Europe, SRLVs are well-known pathogens, but so far, few studies have investigated the circulating viral strains and their prevalence in the islands and mainland. The results of this research highlighted the presence of both A and B subtypes as well as the predominance of a novel A viral cluster. In many cases, the combined serological–molecular approach allowed us to identify more than one viral subtype in a single farm. Small ruminant lentiviruses are a group of viruses infecting goat and sheep worldwide. These viruses exhibit an extraordinary degree of genetic and antigenic variability that severely influence in vivo and in vitro features, as well as diagnostic test results. Small ruminant farming is the most important animal farming business in Greece, with a high impact on the Greek primary economy. Although SRLV infection and its impact on animal production are well established in the country, little is known about the circulating SRLV strains and their prevalence. The aim of this study was to characterize SRLVs circulating in Greece with a combined serological and molecular approach, using the bulk milk matrix collected from 60 farms in different municipalities. This study allowed us to estimate a seroprevalence of around 52% at the herd level. The B1, B2 and A3 subtypes and a novel A viral cluster were identified. Moreover, the amplicon sequencing method allowed us to identify more than one viral subtype in a sample. These results again confirm the high variability of these viruses and highlight the importance of the constant monitoring of viral evolution, in particular in antigens of diagnostic interest. [ABSTRACT FROM AUTHOR]

Urinary tract infections (UTIs) are the most common infections caused mainly by Escherichia coli and the O-antigen and FimH & PapGII adhesion genes are among the most important virulence factors. The aim of this study was to investigate the prevalence of some O-antigen in uropathogenic E. coli (UPEC) and molecular characterization of the adhesion genes FimH & PapGII63. E. coli isolates (50.4% from total number of samples) were collected from urinary tract infection patients from July to October in 2021 and identified using molecular and biochemical tests. Sevenantigenic types of uropathogenic E. coli were investigated, as well as the presence of FimH & PapGII genes in uropathogenic E. coli strains. The results indicated that the majority of UPEC belonged to serotypes O1, O2, O4, O6, O7, O18ac and O75 in varying proportions, where the O1 serotype was the most predominant, followed by O2, O6, O75, O7, O4 and O18ac respectively. The results of molecular tests showed the presence of the FimH gene in 52 isolates with a percentage of 82.50% and the presence of the PapGII gene in 19 isolates with a percentage of 30.15%. From the results we can conclude that the serotypes O1 and O2 had the most virulence role due to presence of the adhesion genes FimH and PapGII. [ABSTRACT FROM AUTHOR]

This narrative review describes genomic characteristic, serotyping, immunogenicity, and vaccine development of Streptococcus pneumoniae capsular polysaccharide (CPS). CPS is a primary virulence factor of S. pneumoniae. The genomic characteristics of S. pneumoniae CPS, including the role of biosynthetic gene and genetic variation within cps (capsule polysaccharide) locus which may lead to serotype replacement are still being investigated. One hundred unique serotypes of S. pneumoniae have been identified through various methods of serotyping using phenotypic and genotypic approach. The advantages and limitations of each method are various, emphasizing the need for accurate and comprehensive serotyping for effective disease surveillance and vaccine targeting. In addition, we elaborate the critical role of CPS in vaccine development by providing an overview of immunogenicity, ongoing research of pneumococcal vaccines, and the impact on disease burden. [ABSTRACT FROM AUTHOR]

Uncommon Salmonella Infantis variants displaying only flagellar antigens phenotypically showed identical incomplete antigenic formula but differed by molecular serotyping. Although most formed rough colonies, all shared antimicrobial resistances and the presence of usg gene with wild-type Salmonella Infantis. Moreover, they were undistinguishable wild-type Salmonella Infantis by whole-genome sequencing. [ABSTRACT FROM AUTHOR]

Background Dengue cases continue to rise and can overwhelm healthcare systems during outbreaks. In dengue, neutrophil mediators, soluble urokinase plasminogen activator receptor (suPAR) and olfactomedin 4, and mast cell mediators, chymase and tryptase, have not been measured longitudinally across the dengue phases. The utility of these proteins as prognostic biomarkers for severe dengue has also not been assessed in an older adult population. Methods We prospectively enrolled 99 adults with dengue—40 dengue fever, 46 dengue with warning signs and 13 severe dengue, along with 30 controls. Plasma levels of suPAR, olfactomedin 4, chymase and tryptase were measured at the febrile, critical and recovery phases in dengue patients. Results The suPAR levels were significantly elevated in severe dengue compared to the other dengue severities and controls in the febrile (P <.001), critical (P <.001), and recovery (P =.005) phases. In the febrile phase, suPAR was a prognostic biomarker of severe dengue, with an AUROC of 0.82. Using a cutoff derived from Youden's index (5.4 ng/mL) and an estimated prevalence of severe dengue (16.5%) in our healthcare institution, the sensitivity was 71.4% with a specificity of 87.9% in the febrile phase, and the positive and negative predictive values were 54.7% and 95.8%, respectively. Olfactomedin 4 was elevated in dengue patients but not in proportion to disease severity in the febrile phase (P =.04) There were no significant differences in chymase and tryptase levels between dengue patients and controls. Conclusions In adult dengue, suPAR may be a reliable prognostic biomarker for severe dengue in the febrile phase. [ABSTRACT FROM AUTHOR]

Background Quantitative polymerase chain reaction (qPCR) targeting ipaH has been proven to be highly efficient in detecting Shigella in clinical samples compared to culture-based methods, which underestimate Shigella burden by 2- to 3-fold. qPCR assays have also been developed for Shigella speciation and serotyping, which is critical for both vaccine development and evaluation. Methods The Enterics for Global Health (EFGH) Shigella surveillance study will utilize a customized real-time PCR–based TaqMan Array Card (TAC) interrogating 82 targets, for the detection and differentiation of Shigella spp, Shigella sonnei , Shigella flexneri serotypes, other diarrhea-associated enteropathogens, and antimicrobial resistance (AMR) genes. Total nucleic acid will be extracted from rectal swabs or stool samples, and assayed on TAC. Quantitative analysis will be performed to determine the likely attribution of Shigella and other particular etiologies of diarrhea using the quantification cycle cutoffs derived from previous studies. The qPCR results will be compared to conventional culture, serotyping, and phenotypic susceptibility approaches in EFGH. Conclusions TAC enables simultaneous detection of diarrheal etiologies, the principal pathogen subtypes, and AMR genes. The high sensitivity of the assay enables more accurate estimation of Shigella -attributed disease burden, which is critical to informing policy and in the design of future clinical trials. [ABSTRACT FROM AUTHOR]

Background: Pneumococcal disease is still considered a global problem. With the introduction of pneumococcal conjugate vaccines (PCVs) serotype epidemiology changed, but antimicrobial resistance persists constituting a serious problem. The current study aimed to determine the serotype distribution and the antimicrobial susceptibility of recent Streptococcus pneumoniae isolates, following implementation of the 13-valent conjugate vaccine (PCV13). Materials and Methods: From January 2017 to December 2022 we evaluated 116 nonduplicate S. pneumoniae isolates collected from adult patients (21 - 98 years) cared for in the University Hospital of Heraklion, Crete, Greece. Pneumococcal isolates were serotyped by the Quellung reaction, and antimicrobial susceptibility testing was performed using E-test. Multidrug resistance (MDR) was defined as non-susceptibility to at least one agent in ≥3 classes of antibiotics. Results: Among the 116 isolates, 31% were recognized as invasive pneumococcal strains, while 69% were noninvasive. The isolates tested belonged to 25 different serotypes. The most prevalent serotypes were 11A (10.3%), and 35B (10.3%), followed by 3 (9.5%), 15A (7.8%), 25F (6.9%), 19A (5.3%), 35F (5.3%), and others (44.6%). The coverage rates of PCV13 and the pneumococcal polysaccharide vaccine (PPSV23) were 26.7% and 57.8%, respectively. PCV13 and PPSV23 serotypes decreased between 2017 - 2019 and 2020 - 2022, with a parallel increase in the non-vaccine types. Resistance rates to erythromycin, clindamycin, trimethoprim/sulfamethoxazole, penicillin, levofloxacin, and ceftriaxone, were 40.5%, 21.6%, 13.8%, 12.1%, 3.4%, and 0%, respectively. All isolates were susceptible to vancomycin, linezolid, and daptomycin. MDR was observed among 36 (31%) S. pneumoniae isolates. Conclusion: The increasing levels of resistance in S. pneumoniae in Crete, Greece, highlight the need for continuous surveillance of antimicrobial resistance and development of strategies for its reduction, including antimicrobial stewardship programs, increased pneumococcal vaccination, and development of next generation PCVs with a wider serotype coverage. [ABSTRACT FROM AUTHOR]

Three hundred samples, including meat from the slaughtered carcass and water, air samples, and swabs from the floor, wall, and employees' hands, were collected from five municipal abattoirs spread across several Egyptian provinces. The Escherichia coli was isolated from floor swabs, meat, air, wall, hand, and water samples. Serotyping of the recovered isolates clarified the presence of various serotypes, including enterohemorrhagic serotypes (O111: H4, O128: H2, and O127: H6) and enterotoxigenic serotypes (O44: H18 and O125: H21). The isolates were resistant to cefotaxime (100%), amoxiclav (80%), then rifampin (66.7%). The stx1 gene, stx2 gene, eaeA gene, blaCMY2 gene and iss gene were detected in 10-80 % of the isolates. Nanosilver (AgNPs) showed that 12.5 ppm was the lowest concentration that prevented bacterial growth. It was observed that 12% of workers wore a clean white coat, only 24% washed their hands between activities during work, only 14% used soap for hand washing, and 42% utilized the same knife for meat and its offal. [ABSTRACT FROM AUTHOR]

Salmonella spp. are important foodborne pathogens often associated with infectious outbreaks caused by consuming contaminated food. Although these microorganisms are widely distributed in several animal production chains, there are few records of their occurrence in aquaculture. This study aimed to evaluate the presence and antimicrobial resistance of Salmonella spp. isolated from fish farms. Samples were collected from three fish farms, covering water, fish, and epilithic biofilm. Salmonella isolates were identified by PCR, and serotyping was performed using the slide agglutination method, according to the Kaufman-White system. Susceptibility to antimicrobial agents was assessed using the Kirby and Bauer disk diffusion method. The presence of class 1 and 2 integrons was determined by PCR. Ninety samples were examined, of which 13 (14.44%) were positive for S. enterica, of which 5 strains were isolated from lagoon water, 3 from fish, and 5 from epilithic biofilm. Among the samples, the serotypes S. Minnesota, S. Panamá, and S. Anatum were identified. Antimicrobial resistance rates were highest for sulfonamides (92.30%), trimethoprim (84.61%), tetracycline (46.15%), and streptomycin (46.15%). Multiple antibiotic resistance was confirmed in 84.61% of the isolates, with 100% presenting integron class 1 and 7.69% integron class 2. This study demonstrated a high prevalence of multiresistant bacteria in fish farms, which reinforces public health concerns due to the use of antibiotics and the risk associated with food from farmed freshwater fish. Given these results, the need for new external research into control and prevention strategies for Salmonella spp. is evident. in aquaculture, as well as greater regulation in fish production chains.

Bacterial pathogenic strains are as adaptable as Salmonella strains and cause diverse intestinal and extraintestinal diseases in humans and other mammals worldwide. Red meat and its products are important hosts for many zoonotic diseases. This work was designed to investigate the frequency, serotypes, and antimicrobial resistance profile of isolated Salmonella spp. in red meat (cattle, sheep, and goats) sold in Dhamar Governorate, Yemen. A total of 250 red meat samples were collected from the retail seller market between July and December 2022. All samples were transported immediately to the laboratory, subcultured on selective enrichment agar, and identified by serotyping and antimicrobial susceptibility tests via disk diffusion methods. The results indicated 26 positive samples of Salmonella out of the 250 samples (10.4%). Notably, isolates belong to ten various serotypes: S. Typhimurium 19.2%, S. Anatum 15.4%, S. Newport 11.5%, S. Enteritidis 11.5%, S. Muenchen 11.5%, S. Infantis 7.7%, S. Montevideo 7.7%, S. Dublin 7.7%, S. Senftenberg 3.9%, and S. Arizona 3.9%. The antibiotic resistance profile revealed that 57.5%, 53.9%, and 53.9% of isolates are resistant to erythromycin, tetracycline, and norfloxacin, respectively. This resistance among Salmonella spp. suggests a significant threat to health, which will in turn require an active safety measure and response. On the other hand, the seasonal variations “August and July” were found to be associated with an increased frequency of Salmonella isolation.

ABSTRACT The International Circumpolar Surveillance (ICS) program is a population-based surveillance network for invasive bacterial diseases throughout Arctic countries and territories. The ICS quality control program for Streptococcus pneumoniae serotyping and antimicrobial susceptibility testing has been ongoing since 1999. Current participating laboratories include the Provincial Laboratory for Public Health in Edmonton, Alberta; Laboratoire de santé publique du Québec in Sainte-Anne-de-Bellevue, Québec; the Centers for Disease Control’s Arctic Investigations Program in Anchorage, Alaska; the Neisseria and Streptococcus Reference Laboratory at Statens Serum Institut in Copenhagen, Denmark; the Department of Clinical Microbiology, Landspitali in Reykjavik, Iceland; and Public Health Agency of Canada’s National Microbiology Laboratory in Winnipeg, Manitoba. From 2009 to 2020, 140 isolates of S. pneumoniae were distributed among the six laboratories as part of the quality control program. Overall serotype concordance was 96.9%, with 99.3% concordance to pool level. All participating laboratories had individual concordance rates >92% for serotype and >97% for pool. Overall concordance by modal minimum inhibitory concentration (MIC) for testing done by broth microdilution or Etest was 99.1%, and >98% for all antimicrobials tested. Categorical concordance was >98% by both CLSI and EUCAST criteria. For two laboratories performing disc diffusion, rates of concordance by modal MIC were >97% for most antimicrobials, except chloramphenicol (>93%) and trimethoprim/sulfamethoxazole (>88%). Data collected from 12 years of the ICS quality control program for S. pneumoniae demonstrate excellent (≥95%) overall concordance for serotype and antimicrobial susceptibility testing results across six laboratories.IMPORTANCEArctic populations experience several social and physical challenges that lead to the increased spread and incidence of invasive diseases. The International Circumpolar Surveillance (ICS) program was developed to monitor five invasive bacterial diseases in Arctic countries and territories. Each ICS organism has a corresponding interlaboratory quality control (QC) program for laboratory-based typing, to ensure the technical precision and accuracy of reference testing services for these regions, and identify and correct potential problems. Here, we describe the results of the ICS Streptococcus pneumoniae QC program, from 2009 to 2020. Excellent overall concordance was achieved for serotype and antimicrobial susceptibility testing results across six laboratories. Ongoing participation in these QC programs ensures the continuation of quality surveillance systems within Arctic populations that experience health disparities.

Abstract Knowing about the antibiotic resistance, serotypes, and virulence-associated genes of Group B Streptococcus for epidemiological and vaccine development is very important. We have determined antimicrobial susceptibility patterns, serotype, and virulence profiles. The antibiotic susceptibility was assessed for a total of 421 Streptococcus agalactiae strains, isolated from pregnant women and neonates. Then, 89 erythromycin and/or clindamycin-resistant strains (82 isolates obtained from pregnant women and seven isolates derived from neonates) were assessed in detail. PCR techniques were used to identify the studied strains, perform serotyping, and assess genes encoding selected virulence factors. Phenotypic and genotypic methods determined the mechanisms of resistance. All tested strains were sensitive to penicillin and levofloxacin. The constitutive MLSB mechanism (78.2%), inducible MLSB mechanism (14.9%), and M phenotype (6.9%) were identified in the macrolide-resistant strains. It was found that macrolide resistance is strongly associated with the presence of the ermB gene and serotype V. FbsA, fbsB, fbsC, scpB, and lmb formed the most recurring pattern of genes among the nine surface proteins whose genes were analysed. A minority (7.9%) of the GBS isolates exhibited resistance to lincosamides and macrolides, or either, including those that comprised the hypervirulent clone ST-17. The representative antibiotic resistance pattern consisted of erythromycin, clindamycin, and tetracycline resistance (71.9%). An increase in the fraction of strains resistant to macrolides and lincosamides indicates the need for monitoring both the susceptibility of these strains and the presence of the ST-17 clone.

Knowing about the antibiotic resistance, serotypes, and virulence-associated genes of Group B Streptococcus for epidemiological and vaccine development is very important. We have determined antimicrobial susceptibility patterns, serotype, and virulence profiles. The antibiotic susceptibility was assessed for a total of 421 Streptococcus agalactiae strains, isolated from pregnant women and neonates. Then, 89 erythromycin and/or clindamycin-resistant strains (82 isolates obtained from pregnant women and seven isolates derived from neonates) were assessed in detail. PCR techniques were used to identify the studied strains, perform serotyping, and assess genes encoding selected virulence factors. Phenotypic and genotypic methods determined the mechanisms of resistance. All tested strains were sensitive to penicillin and levofloxacin. The constitutive MLSB mechanism (78.2%), inducible MLSB mechanism (14.9%), and M phenotype (6.9%) were identified in the macrolide-resistant strains. It was found that macrolide resistance is strongly associated with the presence of the ermB gene and serotype V. FbsA, fbsB, fbsC, scpB, and lmb formed the most recurring pattern of genes among the nine surface proteins whose genes were analysed. A minority (7.9%) of the GBS isolates exhibited resistance to lincosamides and macrolides, or either, including those that comprised the hypervirulent clone ST-17. The representative antibiotic resistance pattern consisted of erythromycin, clindamycin, and tetracycline resistance (71.9%). An increase in the fraction of strains resistant to macrolides and lincosamides indicates the need for monitoring both the susceptibility of these strains and the presence of the ST-17 clone. [ABSTRACT FROM AUTHOR]

Simple Summary: The characterization of small ruminant lentiviruses in autochthonous goat and sheep populations, like the Moroccan one, represents an excellent source of information to understand their evolution from domestication to present day and improve the knowledge of the genetic diversity of these viruses. Serological data confirmed a low prevalence likely due to extensive breeding and management practice in this area. However, the low rate of success of routinely used serological tests led us to suppose that divergent strains might have escaped diagnostic tools thanks to mutations' diagnostic epitopes. This hypothesis is confirmed by next-generation sequencing results, which highlighted the presence of an A subtype carrying a mismatch in the serotyping epitope. Moreover, the circulation of novel B and recombinant A/B subtypes was also revealed. Recent studies that investigated the origins of SRLV strains offered new insights into their distribution among domestic ruminants. The aim of the study was to investigate SRLV circulation in Morocco. A total of 51 farms were selected in different geographical locations and tested by screening and genotyping ELISA. Whole blood was used for DNA extraction and nested gag PCR. The sample size allowed for an estimation of prevalence lower than 20% (CI 95%). Surprisingly, a large proportion of screening-positive samples were not correctly serotyped. Sanger and NGS amplicon sequencing approaches allowed us to obtain new sequences even from difficult-to-amplify samples. The serological data support the evidence of an intrinsic difficulty of SRLV to spread, likely due to management practices. The low rate of success by genotyping ELISA led us to suppose that divergent strains might have escaped from diagnostic tools, as partially confirmed by the evidence of an A subtype carrying a mismatch in serotyping epitope. The sequence analysis revealed the circulation of novel B and recombinant A/B subtypes. This study highlights the importance of monitoring viral sequences and their evolution to develop specific diagnostic tests, particularly in countries where control measures are in place. [ABSTRACT FROM AUTHOR]

OBJECTIVES: We sought to describe the evolving epidemiology of invasive pneumococcal disease (IPD) among children in Massachusetts, United States, over the last 2 decades during which sequential 7-valent pneumococcal conjugate vaccines (PCV7) and 13-valent PCVs (PCV13) were implemented. METHODS: Cases of IPD in children aged <18 years were detected between 2002 and 2021 through an enhanced population-based, statewide surveillance system. Streptococcus pneumoniae isolates from normally sterile sites were serotyped and evaluated for antimicrobial susceptibility. IPD incidence rates and rate ratios with 95% confidence intervals (CIs) were calculated. RESULTS: We identified 1347 IPD cases. Incidence of IPD in children aged <18 years declined 72% over 2 decades between 2002 and 2021 (incidence rate ratios 0.28, 95% CI 0.18--0.45). IPD rates continued to decline after replacement of PCV7 with PCV13 (incidence rate ratios 0.25, 95% CI 0.16--0.39, late PCV7 era [2010] versus late PCV13 era [2021]). During the coronavirus disease 2019 pandemic years, 2020 to 2021, the rate of IPD among children aged <18 years reached 1.6 per 100 000, the lowest incidence observed over the 20 years. In PCV13 era, approximately one-third of the IPD cases in children aged >5 years had at least 1 underlying condition (98, 30.3%). Serotypes 19A and 7F contributed 342 (48.9%) of all cases before implementation of PCV13 (2002--2010). Serotype 3 (31, 8.6%), and non-PCV13 serotypes 15B/C (39, 10.8%), 33F (29, 8.0%), 23B (21, 0.8%), and 35B (17, 4.7%) were responsible for 37.8% of cases in PCV13 era (2011--2021). Penicillin nonsusceptibility continued to decline (9.8% vs 5.3% in pre-/late PCV13 era, P 5 .003), however has become more common among non-PCV13 serotypes compared with vaccine serotypes (14.8% vs 1.4%, P < .001). CONCLUSIONS: Robust ongoing surveillance networks are critical for identifying emerging serotypes and development of next-generation vaccine formulations. [ABSTRACT FROM AUTHOR]

Hong Kong is an Asia-Pacific City with low incidence but periodic local outbreaks of dengue. A mixed-method assessment of the risk of expansion of dengue endemicity in such setting was conducted. Archived blood samples of healthy adult blood donors were tested for anti-dengue virus IgG at 2 time-points of 2014 and 2018/2019. Data on the monthly notified dengue cases, meteorological and vector (ovitrap index) variables were collected. The dengue virus (DENV) IgG seroprevalence of healthy adults in 2014 was 2.2% (95%C.I. = 1.8–2.8%, n = 3827) whereas that in 2018/2019 was 1.7% (95%C.I. = 1.2–2.3%, n = 2320). Serotyping on 42 sera in 2018/2019 showed that 22 (52.4%) were DENV-2. In 2002–2019, importation accounted for 95.3% of all reported cases. By wavelet analysis, local cases were in weak or no association with meteorological and vector variables. Without strong association between local cases and meteorological/vector variables, there was no evidence of increasing level of dengue infection in Hong Kong. [ABSTRACT FROM AUTHOR]

Background and Aim: Non-Typhoidal Salmonellae (NTS) is emerging as a major health problem globally. They cause foodborne infections, especially gastroenteritis as well as extraintestinal manifestations. As this is a zoonotic disease, certain animal sources such as poultry, egg, meat and dairy products are implicated. Fruits and vegetables too contaminated by manure of infected animal excreta are possible vehicles of transmission of this disease. The present report analyzes the spectrum of NTS with seroprofiling, isolated in our institute for the last 12 years. Materials and Methods: Fresh fecal samples from patients with acute gastroenteritis and blood samples from cases of bacteremia were processed according to the standard protocol. Nonlactose fermenting colonies resembling Salmonella were subjected to biochemical characterization and antimicrobial susceptibility testing followed by serotyping. Results: One hundred fifty-two NTS strains were isolated from feces and 13 from blood. Serotyping of these strains revealed that the most common NTS species was Salmonella Typhimurium (44.8%), followed by Salmonella Oslo (15.2%) and Salmonella Weltevreden (11.5%). Many rare strains were also encountered in our study. Conclusion: As many of the NTS species are biochemically similar, serotyping of NTS should be made mandatory to know the burden of the disease and to detect the emergence of new serovars. Emergence of multidrug resistance in NTS strains is a major concern nowadays. It is very essential to adopt strict quality measures in poultry, fish, dairy, and meat processing units as they are widely distributed in animals. In addition, strategies should be formulated by health education and public awareness programs to sensitize the population regarding this menace. [ABSTRACT FROM AUTHOR]

Gulls act as intermediaries in the exchange of microorganisms between the environment and human settlements, including Salmonella spp. This study assessed the antimicrobial resistance and molecular profiles of Salmonella spp. isolates obtained from fecal samples of gulls in the city of Porto, Portugal, in 2008 and 2023 and from water samples in 2023. Antimicrobial susceptibility profiling revealed an improvement in the prevalence (71% to 17%) and antimicrobial resistance between the two collection dates. Two isolate collections from both 2008 and 2023 underwent serotyping and whole-genome sequencing, revealing genotypic changes, including an increased frequency in the monophasic variant of S. Typhimurium. qacE was identified in 2008 and 2023 in both water and fecal samples, with most isolates exhibiting an MDR profile. The most frequently observed plasmid types were IncF in 2008 (23%), while IncQ1 predominated in 2023 (43%). Findings suggest that Salmonella spp. circulate between humans, animals, and the environment. However, the genetic heterogeneity among the isolates from the gulls' feces and the surface water may indicate a complex ecological and evolutionary dynamic shaped by changing conditions. The observed improvements are likely due to measures to reduce biological contamination and antimicrobial resistance. Nevertheless, additional strategies must be implemented to reduce the public health risk modeled by the dissemination of pathogens by gulls. [ABSTRACT FROM AUTHOR]

Introduction: Whole Genome Sequencing (WGS) implementation in food safety laboratories is a significant advancement in food pathogen control and outbreak tracking. However, the initial investment for acquiring next-generation sequencing platforms and the need for bioinformatic skills represented an obstacle for the widespread use of WGS. Long-reading technologies, such as the one developed by Oxford Nanopore Technologies, can be easily implemented with a minor initial investment and with simple protocols that can be performed with basic laboratory equipment. Methods: Herein, we report a simple MinION Galaxy-based workflow with analysis parameters that allow its implementation in food safety laboratories with limited computer resources and without previous knowledge in bioinformatics for rapid Salmonella serotyping, virulence, and identification of antimicrobial resistance genes. For that purpose, the single use Flongle flow cells, along with the MinION Mk1B for WGS, and the community-driven web-based analysis platform Galaxy for bioinformatic analysis was used. Three strains belonging to three different serotypes, monophasic S. Typhimurium, S. Grancanaria, and S. Senftenberg, were sequenced. Results: After 24 h of sequencing, enough coverage was achieved in order to perform de novo assembly in all three strains. After evaluating different tools, Flye de novo assemblies with medaka polishing were shown to be optimal for in silico Salmonella spp. serotyping with SISRT tool followed by antimicrobial and virulence gene identification with ABRicate. Discussion: The implementation of the present workflow in food safety laboratories with limited computer resources allows a rapid characterization of Salmonella spp. isolates. [ABSTRACT FROM AUTHOR]

Haemophilus influenzae is one of the main bacteria responsible for otitis media (OM) among children worldwide. We aimed to estimate the distribution of encapsulated and non-capsulated variants (NTHi), biotypes, antibiotic susceptibility, and molecular epidemiology of H. influenzae isolates recovered from pediatric OM cases in Bulgaria. Capsule detection was done by PCR for bexB gene, absent in NTHi. All encapsulated strains were subjected to PCR serotyping. MIC susceptibility testing was performed according to the criteria of EUCAST. MLST was conducted for all 71 OM isolates. The capsule detection and PCR – serotyping disclosed a predominance of NTHi (90.1%) and a few "a", "f", and "c" types. Biotype I was the most widespread (42.3%). β-lactam resistance was found in 35.2% of the isolates. MLST represented heterogenic population structure, whereas the most represented clonal complexes belonged to ST-3, ST-57, ST-105, and ST-1426. 42.3% of the STs showed relatedness to globally represented clones, and 11.3% displayed affiliation to international type 2. Most of the H. influenzae isolates recovered from children with otitis media were non-typable strains from biotype I. The examined population structure was genetically diverse, with a predominance of international type 2 isolates. [ABSTRACT FROM AUTHOR]

The clinical consequences of toxoplasmosis are greatly dependent on the Toxoplasma gondii strain causing the infection. To better understand its epidemiology and design appropriate control strategies, it is important to determine the strain present in infected animals. Serotyping methods are based on the detection of antibodies that react against segments of antigenic proteins presenting strain-specific polymorphic variations, offering a cost-effective, sensitive, and non-invasive alternative to genotyping techniques. Herein, we evaluated the applicability of a panel of peptides previously characterized in mice and humans to serotype sheep and pigs. To this end, we used 51 serum samples from experimentally infected ewes (32 type II and 19 type III), 20 sheep samples from naturally infected sheep where the causative strain was genotyped (18 type II and 2 type III), and 40 serum samples from experimentally infected pigs (22 type II and 18 type III). Our ELISA test results showed that a combination of GRA peptide homologous pairs can discriminate infections caused by type II and III strains of T. gondii in sheep and pigs. Namely, the GRA3-I/III-43 vs. GRA3-II-43, GRA6-I/III-213 vs. GRA6-II-214 and GRA6-III-44 vs. GRA6-II-44 ratios showed a statistically significant predominance of the respective strain-type peptide in sheep, while in pigs, in addition to these three peptide pairs, GRA7-II-224 vs. GRA7-III-224 also showed promising results. Notably, the GRA6-44 pair, which was previously deemed inefficient in mice and humans, showed a high prediction capacity, especially in sheep. By contrast, GRA5-38 peptides failed to correctly predict the strain type in most sheep and pig samples, underpinning the notion that individual standardization is needed for each animal species. Finally, we recommend analyzing for each animal at least 2 samples taken at different time points to confirm the obtained results.

In this study, an automated, targeted next-generation sequencing (tNGS) assay to detect and serotype Salmonella from sample enrichments was evaluated. The assay generates millions of reads to detect multiple Salmonella-specific genes and serotype-specific alleles, detecting all Salmonella spp. tested to date, and serotyping 62 common Salmonella serotypes. Accuracy was tested on 291 pure reference cultures (251 Salmonella, 40 non-Salmonella), 21 artificially contaminated poultry carcass rinse samples, and 363 naturally contaminated poultry environmental samples. Among the 291 pure reference cultures, the automated tNGS assay resulted in 100% detection accuracy, 100% serotyping accuracy for the claimed serotypes, and 0% false positives. The limit of detection was estimated at 5 × 104 CFU/mL by testing enumerated cultures of strains representative of six serotypes. In cocontamination studies with mixtures of two serotypes (Enteritidis, Typhimurium, Kentucky, Infantis, and Newport) at a 1:1 ratio, tNGS detected both serotypes with 100% accuracy. The assay demonstrated 100% accuracy in artificially contaminated poultry carcass rinse sample enrichments. Targeted NGS was highly effective in detecting Salmonella in samples collected from poultry production facilities. Results demonstrated that tNGS could detect Salmonella and provide accurate serotyping information consistent with conventional serology. These findings highlight the reliable and efficient performance of a fully automated tNGS Salmonella assay in detecting and identifying Salmonella strains in complex matrices, reducing the time to results from 4 to 5 days required by the traditional isolation and serotyping to 10–12 h for tNGS after primary enrichment.

ABSTRACTThe resolution of variation within species is critical for interpreting and acting on many microbial measurements. In the key foodborne pathogens Salmonella and Escherichia coli, the primary subspecies classification scheme used is serotyping: differentiating variants within these species by surface antigen profiles. Serotype prediction from whole-genome sequencing (WGS) of isolates is now seen as comparable or preferable to traditional laboratory methods where WGS is available. However, laboratory and WGS methods depend on an isolation step that is time-consuming and incompletely represents the sample when multiple strains are present. Community sequencing approaches that skip the isolation step are, therefore, of interest for pathogen surveillance. Here, we evaluated the viability of amplicon sequencing of the full-length 16S rRNA gene for serotyping Salmonella enterica and E. coli. We developed a novel algorithm for serotype prediction, implemented as an R package (Seroplacer), which takes as input full-length 16S rRNA gene sequences and outputs serovar predictions after phylogenetic placement into a reference phylogeny. We achieved over 89% accuracy in predicting Salmonella serotypes on in silico test data and identified key pathogenic serovars of Salmonella and E. coli in isolate and environmental test samples. Although serotype prediction from 16S rRNA gene sequences is not as accurate as serotype prediction from WGS of isolates, the potential to identify dangerous serovars directly from amplicon sequencing of environmental samples is intriguing for pathogen surveillance. The capabilities developed here are also broadly relevant to other applications where intraspecies variation and direct sequencing from environmental samples could be valuable.IMPORTANCEIn order to prevent and stop outbreaks of foodborne pathogens, it is important that we can detect when pathogenic bacteria are present in a food or food-associated site and identify connections between specific pathogenic bacteria present in different samples. In this work, we develop a new computational technology that allows the important foodborne pathogens Escherichia coli and Salmonella enterica to be serotyped (a subspecies level classification) from sequencing of a single-marker gene, and the 16S rRNA gene often used to surveil bacterial communities. Our results suggest current limitations to serotyping from 16S rRNA gene sequencing alone but set the stage for further progress that we consider likely given the rapid advance in the long-read sequencing technologies and genomic databases our work leverages. If this research direction succeeds, it could enable better detection of foodborne pathogens before they reach the public and speed the resolution of foodborne pathogen outbreaks.

Abstract Objective Determining the serotype of circulating virus strains is important in implementing effective vaccination. In this study, Foot-and-Mouth Disease (FMD) Southern African territory 2 (SAT2) specific primers and TaqMan probe were designed towards rapid SAT2 detection and serotyping. The primers were tested by endpoint reverse transcription (RT) polymerase chain reaction (PCR) and quantitative PCR (RT-qPCR) using the vaccine strain SAT2035. The SAT2 serotype-specific RT-qPCR assay was compared with currently used ELISA and VP1 sequencing using Cohen’s kappa statistics. Results The primers yielded amplicons of band size 190 bp during endpoint RT-PCR. When coupled with the probe, the primers reaction efficiency was determined to be 99% with an r2 value of 0.994. The results show that the SAT2 assay has comparable performance to VP1 sequencing (k = 1) and a moderate degree of agreement with ELISA (k = 0.571). The data shows that the newly designed assay could be considered for serotyping of SAT2 strains. However, for this assay to be complete there is a need to design effective SAT1 and SAT3 primers and probes that can be multiplexed to target other serotypes that co-circulate within relevant FMD endemic pools. For future implementation of the assay there is also a need to increase the number of field samples towards validation of the assay.