Agalactiae Isolates Research Articles

Comprehensive Characterization of a Streptococcus agalactiae Phage Isolated from a Tilapia Farm in Selangor, Malaysia, and Its Potential for Phage Therapy.

The Streptococcus agalactiae outbreak in tilapia has caused huge losses in the aquaculture industry worldwide. In Malaysia, several studies have reported the isolation of S. agalactiae, but no study has reported the isolation of S. agalactiae phages from tilapia or from the culture pond. Here, the isolation of the S. agalactiae phage from infected tilapia is reported and it is named as vB_Sags-UPM1. Transmission electron micrograph (TEM) revealed that this phage showed characteristics of a Siphoviridae and it was able to kill two local S. agalactiae isolates, which were S. agalactiae smyh01 and smyh02. Whole genome sequencing (WGS) of the phage DNA showed that it contained 42,999 base pairs with 36.80% GC content. Bioinformatics analysis predicted that this phage shared an identity with the S. agalactiae S73 chromosome as well as several other strains of S. agalactiae, presumably due to prophages carried by these hosts, and it encodes integrase, which suggests that it was a temperate phage. The endolysin of vB_Sags-UPM1 termed Lys60 showed killing activity on both S. agalactiae strains with varying efficacy. The discovery of the S. agalactiae temperate phage and its antimicrobial genes could open a new window for the development of antimicrobials to treat S. agalactiae infection.

Molecular characteristics and antibiotic resistance mechanisms of clindamycin-resistant Streptococcus agalactiae isolates in China.

Streptococcus agalactiae (Group B Streptococcus, GBS) is a major cause of neonatal infections with high morbidity and mortality, and clindamycin is the main antibiotic used to treat GBS infections in patients allergic to penicillin. We aimed to analyse the antibiotic sensitivity, sequence types, serotypes, virulence factors, and antibiotic resistance mechanisms of clinically isolated clindamycin-resistant S. agalactiae and provide basic data for the treatment, prevention, and control of clinical infection of S. agalactiae. A total of 110 strains of clindamycin-resistant S. agalactiae were collected from two tertiary hospitals in Hebei, China. We performed antibiotic sensitivity tests for 11 antibiotics on these strains and whole-genome sequencing analysis. All the strains were susceptible to penicillin, ampicillin, linezolid, vancomycin, tigecycline, and quinupristin-dalfopristin. Resistance to erythromycin, levofloxacin, tetracycline, and chloramphenicol were also observed. Genome sequence analysis revealed that all strains belonged to 12 sequence types (STs) related to six cloning complexes (CCs), namely CC10, CC19, CC23, CC651, CC1, and CC17. Five serotypes were identified, including IA, IB, II, III, and V. The most prominent resistance genes were mreA (100%) and ermB (81.8%). Furthermore, cfb, cylE, pavA and the gene cluster related to the pili were 100% present in all strains, followed by lmb (95.5%) and srr1 (67.2%). This study found that clindamycin-resistant S. agalactiae showed polymorphisms in molecular types and serotypes. Furthermore, multiple virulence factor genes have been identified in their genomes.

Genomic characterization of beta-haemolytic streptococci isolated from prosthetic joint infections.

Prosthetic joint infection (PJI) is an increasing concern for the medical profession, as higher numbers of arthroplasty surgeries lead to rising PJI-related costs. Streptococcal PJIs constitute approximately 10% of PJIs, but their genetic features and characteristics remain largely unexplored. Little is known about antimicrobial resistance (AMR) rates, whether some sequence types (ST) dominate, and whether certain virulence-associated genes are overrepresented. We used whole-genome sequencing of Streptococcus dysgalactiae (n=22), Streptococcus agalactiae (n=10) and S.pyogenes (n=1) to elicit genomic data on 33 beta-haemolytic streptococci isolated from PJIs in Region Örebro county, Sweden. Relatedness was inferred based on single nucleotide polymorphisms in S.dysgalactiae and S.agalactiae. The genomic data were screened for virulence-associated genes available in the Virulence Factor Database. All isolates were screened for both phenotypic and genotypic resistance. The S.dysgalactiae and S.agalactiae isolates were genetically diverse, although 32% of S.dysgalactiae isolates (n=7) were ST20. The speS and PI-2A genes were less represented in these isolates among virulence-associated genes, and AMR was more frequently observed in S.agalactiae. We conclude that PJIs caused by beta-haemolytic streptococci are not dominated by genetically similar beta-haemolytic streptococci. There were distinct inter-species differences in AMR between S.agalactiae and S.dysgalactiae.

Antibiotic resistance genes and molecular typing of Streptococcus agalactiae isolated from pregnant women

BackgroundThe antibiotic resistance of genital tract colonizing Streptococcus agalactiae in pregnant women is increasing. We aimed to determine the antibiotic resistance genes of different clonal types of this bacterium in pregnant women.MethodsFour hundred twenty non-repeated vaginal and rectal specimens were collected from pregnant women and were transferred to the laboratory using Todd Hewitt Broth. The samples were cultured on a selective medium, and the grown bacteria were identified by standard microbiological and biochemical tests. Antimicrobial resistance pattern and inducible clindamycin resistance of the isolates were determined using the disk agar diffusion method. The genomic DNAs of S. agalactiae strains were extracted using an extraction kit, and the antibiotic resistance genes and RAPD types were detected using the PCR method.ResultsThe average age of the participants was 30.74 ± 5.25 years. There was a significant relationship between the weeks of pregnancy and the number of positive bacterial cultures (P-value < 0.05). Moreover, 31 pregnant women had a history of abortion, and 18 had a history of membrane rupture. Among 420 specimens, 106 S. agalactiae isolates were detected. The highest antibiotic resistance rate was found against tetracycline (94.33%), and all isolates were susceptible to linezolid. Moreover, 15, 15, 42, and 7 isolates showed an iMLSB, M-, cMLSB, and L-phenotype. The ermB was the most prevalent resistance gene in the present study, while 38 (35.84%), 8 (7.54%), 79 (74.52%), 37 (34.9%), and 20 (18.86%) isolates were contained the ermTR, mefA/E, tetM, tetO, and aphA3 gene, respectively.ConclusionsThe high-level antibiotic resistance and prevalence of resistance genes may be due to the arbitrarily use, livestock industry consumption, and the preventive use of antibiotics in pregnant women. Thus, the need to re-considering this problem seems to be necessary.

Molecular epidemiology, drug resistance, and virulence gene analysis of Streptococcus agalactiae isolates from dairy goats in backyard farms in China.

Streptococcus agalactiae infections may lead to clinical or subclinical mastitis in dairy animals when it invades the mammary gland. In this study, 51 S. agalactiae strains were isolated from 305 milk samples that were collected from goats with mastitis in 13 provinces of China. The antimicrobial resistance of S. agalactiae was determined by disk diffusion methods against 18 antibiotics from six classes. In addition, multilocus sequence typing (MLST), and the presence of resistance and virulence genes was determined by PCR analysis. Seven sequence types in five clonal complexes were identified according to MLST; CC103 and CC67 strains were predominant, with rates of 45.1% and 39.2%, respectively. All isolates (100%) were multiresistant to three or more antimicrobial agents. S. agalactiae isolates had a 100% resistance rate to penicillin, oxacillin, and amoxicillin, followed by doxycycline (82.4%), tetracycline (76.5%), and amikacin (74.5%). The lowest resistance was observed for ciprofloxacin (29.4%), which varied in five different regions. The detection rates of six classes of antimicrobial-related genes were calculated as follows: 33 (64.7%) for β-lactam-related resistance gene, 12 (23.5%) for tetracyclines, 11 (21.6%) for quinolone-related resistance genes, 10 (19.6%) for aminoglycosides, 7 (13.7%) for macrolides (ermA, ermB, and mefA), and 3 (5.9%) for lincosamide (lnu(B)). Regarding virulence genes, profile 1 (bca cfb-cspA-cylE-hylB-bibA-pavA-fbsA-fbsB) was the most prevalent, with a detection rate of 54.9%. This work provides a primary source related to the molecular epidemiology of S. agalactiae in dairy goat herds in China and will aid in the clinical treatment, prevention, and control of mastitis.

Characteristics of Streptococcus agalactiae causing urinary tract infections: Emergence of new sequence types ST74 and ST38 in Iran.

Difficult-to-treat Streptococcus agalactiae infections are increasingly described in patients with urinary tract infections (UTIs). This occurrence could be due to the production of virulence determinants. This study aimed to characterize the molecular features of S. agalactiae responsible for UTIs. In this cross-sectional study, 70 S. agalactiae isolated from UTIs were examined. Antibiotic susceptibility testing was performed using the disk diffusion method. All S. agalactiae isolates were confirmed by atr and dltS PCR assays. Virulence, alpha protein-like, and pilus island genes were detected by PCR. Isolates were characterized using the multilocus sequence typing method. Multidrug resistance was observed in 80% of isolates. Five virulence profiles were detected, wherein cylE, lmb, bca, rib (35.7%), cylE, lmb, alp3 (27.1%), and cylE, lmb, bac, rib, alp2 (21.4%) were the most frequent detected profiles. S. agalactiae was isolated and categorized within three clonal complexes (CCs) including CC22 (40%), CC17 (25.7%), and CC23 (20%). The main sequence types (STs) found were ST22 (27.1%), ST23 (17.1%), ST17 (12.9%), ST31 (8.7%), ST40 (8.7%), ST74 (7.1%), ST48 (4.3%), ST890 (4.3%), ST189 (2.8%), ST38 (2.8%), ST52 (2.8%), and ST155 (1.4%). ST74 and ST38 were reported for the first time in Tehran-Iran. This study highlights the predominance of the CC22 lineage among S. agalactiae strains isolated from UTIs in Tehran, Iran, and highlights the significant penetration of this lineage into hospitals. MDR patterns among these strains appear to be becoming a major concern in the management of infections.

Prevalence and antimicrobial resistance in Streptococcus agalactiae isolated from raw milk in Pasuruan and Lumajang districts, East Java, Indonesia

Abstract. Permatasari DA, Anggraeni F, Budiarto, Meles DK, Hamid IS, Puspitasari Y, Effendi MH, Khairullah AR, Wardhana DK, Ugbo EN. 2022. Prevalence and antimicrobial resistance in Streptococcus agalactiae isolated from raw milk in Pasuruan and Lumajang districts, East Java, Indonesia. Biodiversitas 23: 5050-5055. Milk and dairy products are nutritionally important in the diet worldwide. The microbiological quality of raw milk is essential for the quality of the final dairy product. The presence of Streptococcus agalactiae is frequently associated with high somatic cell counts in milk and decreased milk yield. Streptococcus agalactiae infections have major consequences for public health. This study aimed to identify the prevalence and antimicrobial resistance of Streptococcus agalactiae isolated from raw milk in Pasuruan and Lumajang, East Java, Indonesia. Raw milk samples were collected from three sub-districts in Pasuruan (Tutur, Grati and Purwosari) and three sub-districts in Lumajang (Senduro, Karangbendo, and Tekung). California mastitis test (CMT) was performed to confirm the presence of somatic cells, S. agalactiae was identified using standard microbiological methods, and antibiotic resistance was determined by Kirby-Bauer disc diffusion method using Mueller-Hinton agar. Results showed that 89 (79.46%) out of 112 dairy cows were positive for CMT, which indicated the presence of mastitis. Bacteria isolation was performed on CMT-positive samples and four samples were showed the presence of S. agalactiae (4.49%). Based on the sensitivity test against various antibiotics, it was observed that S. agalactiae isolates were resistant to ampicillin (75%), and erythromycin (50%), and were sensitive to cefotaxime (100%), tetracycline (75%), and chloramphenicol (100%). This research reported that Streptococcus agalactiae organism is one bacterium that is implicated in mastitis infections. Therefore, monitoring the antibiotic susceptibility profiles of this organism is needed in veterinary medicine, in order to make an appropriate choice of drug for treatment, improve high cure rates and minimize the increase of drug resistance.

Pharmacodynamics of Flucloxacillin in a Neutropenic Murine Thigh Infection Model: A Piece of the Puzzle towards Evidence-Based Dosing.

For decades, flucloxacillin has been used to treat methicillin-susceptible Staphylococcus aureus (MSSA). Little is still known about its pharmacodynamics (PD). The present study aimed to determine the pharmacokinetic (PK)/PD index and the PD-index value minimally required for efficacy. MICs of 305 MSSA isolates were measured to determine the wild-type distribution. The PD of 8 S. aureus, 1 S. pyogenes, and 1 S. agalactiae isolates were evaluated in a neutropenic murine thigh infection model. Two S. aureus isolates were used in a dose-fractionation study and a dose–response analysis was performed additionally in the in vivo model. Data were analyzed with a population PK and sigmoid maximum effect model. The end of the wild-type distribution was 1 mg/L. The percentage of time the unbound concentration was above MIC (%fT > MIC) was best correlated with efficacy. For S. aureus, median %fT > 0.25 × MIC required for 1-log reduction was 15%. The value for S. pyogenes was 10%fT > MIC and for S. agalactiae 22%fT > 0.25xMIC for a 1-log reduction. The effect of flucloxacillin reached a 2-log reduction of S. aureus at 20%fT > 0.25xMIC and also for S. pyogenes and S. agalactiae, a reduction was reached. These data may serve to optimize dosing regimens currently used in humans.

Isolation and identification of streptococcus agalactiae colonization in pregnancy woman and resistance to tetracycline in Al_Muthanna proviance

S.agalactiae is a Gram-positive, spherical bacteria that causes postpartum sepsis, endometritis, chorioamnionitis, and premature birth in pregnant women.Streptococcus agalactiae, often known as group B streptococci (GBS), is a dangerous disease that can infect both humans and animals (GBS) is an important bacterial pathogen that causes a wide range of infections including eonatal sepsis, meningitis, pneumonia and soft tissue or urinary tract infections.The present study demonstrated that S. agalactiae isolates obtained from vaginal samples showed a high rate of resistance to tetracycline. Objectives: The study aimed to investigate streptococcus aqalctaein pregnancy woman by isolating and diagnosing them from rectovaginal colonization .Result We were collected 206 swab from pregnant female this collected was during gestation period (35-37)week and were collected from different age that including (≤25 ) years,and (≥25years .this sample collected from different hospital included (AL_Rumaitha Hospital, womans and childrens hospital ).the collected swab had transfer swabs by Amis transported medium and give to lab. After that diagnosis by microscobic and biochemical test. S. agalactiae was spread through pregnant female and infection the infant.

Antibiotic Sensitivity Profile of Mastitis Pathogens from Dairy Cows in a Selected Farm in Laguna, Philippines [RESEARCH NOTE

Due to their inappropriate and indiscriminate use in treating mastitis, most antibiotics are now ineffective. The aim of the study was to determine the antibiotic sensitivity profile of identified bovine mastitis pathogens. Twenty-seven out of 38 isolates were revived from a previous work and were identified for antibiotic sensitivity. Gram-negative isolates were Klebsiella pneumoniae, Brevundimonas diminuta, Escherichia coli, Acinetobacter baumanii, Vibrio metschnikovii, Flavimonas oryzihabitans and Pantoea agglomerans. Gram-positive isolates were Streptococcus agalactiae, Lactococcus lactis, Streptococcus sp. group C or G, and Corynebacterium renale. Enterobacteriaceae isolates were resistant to Beta lactams, Erythromycin, Nitrofurantoin, Novobiocin, and Clindamycin but were sensitive to Fluroquinones. Miscellaneous Gram-negative isolates were sensitive to Colistin and Ofloxacin but were resistant to Beta lactams, Novobiocin, and Clindamycin. Str. agalactiae isolates were sensitive to Cefaclor, Ofloxacin, and Gentamicin but were resistant to Cloxacillin and Trimethoprim. Gram-positive isolates were sensitive to Oxytetracycline, Ofloxacin, Erythromycin, and Gentamicin. Mastitis treatment guided by antibiotic sensitivity testing is a promising course of therapy likely to succeed. The regimen can be further improved by a combination of non-antibiotic therapeutic methods, responsible use of suitable antibiotics, good sanitation protocols, and organized recording of data.

Pharmacokinetics and pharmacokinetic/pharmacodynamic-based dosing regimens of long-acting oxytetracycline in Nile tilapia (Oreochromis niloticus) broodstock to minimize selection of drug resistance

This study aimed to determine the pharmacokinetics of long-acting oxytetracycline (OTC-LA) after intraperitoneal (IP) administration in Nile tilapia broodstock. Pharmacokinetic/pharmacodynamic (PK/PD) relationships were evaluated to determine the optimal dosing regimens of OTC-LA against Streptococcus agalactiae. A total of 120 healthy male tilapia (450 ± 37.47 g) were arbitrarily divided into two groups (60 fish/group) that individually received OTC-LA single IP injection at dosages of 50 or 100 mg/kg body weight. Blood samples were collected at various times posttreatment, and plasma oxytetracycline (OTC) was analyzed using high-performance liquid chromatography. For pharmacodynamic study, 56 S. agalactiae isolates were subjected to agar dilution to determine the minimum inhibitory concentration (MIC) and mutant prevention concentration (MPC). Results showed that the Cmax and Tmax of OTC-LA were both 110.70 ± 5.61 μg/mL at 2 h for the dosage of 50 mg/kg and 287.85 ± 8.03 μg/mL at 4 h for the dosage of 100 mg/kg. Plasma OTC levels remained at 3.99 ± 0.48 and 23.00 ± 2.51 μg/mL at 168 h (7 day) after OTC-LA administration at 50 and 100 mg/kg, respectively. The MIC50 and MIC90 of S. agalactiae were 0.5 and 1 μg/mL, respectively, and its MPC50 and MPC90 were 32 and 128 μg/mL, respectively. The mutant selection window of MIC50 and MPC50 was 0.5–32 μg/mL, and that of MIC90 and MPC90 was 1–128 μg/mL. OTC-LA at 50 and 100 mg/kg provided plasma OTC levels above MIC for at least 7 days. However, only OTC-LA at 100 mg/kg can provide plasma OTC to prevent the growth of resistant mutant subpopulation. Our study suggested that a single IP administration of OTC-LA at 100 mg/kg body weight is the optimal dosing regimen to attain therapeutic efficacy against S. agalactiae infection in tilapia and prevent the emergence of resistant mutant subpopulation.

Omadacycline Efficacy against Streptococcus Agalactiae Isolated in China: Correlation between Resistance and Virulence Gene and Biofilm Formation.

This study aimed to evaluate the activity, resistance, clonality of MIC distribution, and the correlation between virulence and resistance genes and biofilm formation of omadacycline (OMC) in clinics for Streptococcus agalactiae isolates from China. 162 isolates were collected retrospectively in China. The S. agalactiae were collected from the body's cervical secretions, wound secretions, ear swabs, secretions, semen, venous blood, cerebrospinal fluid, pee, etc. The MIC of OMC against S. agalactiae was determined by broth microdilution. The inhibition zone diameters of OMC and other common antibiotics were measured using filter paper. D-test was performed to determine the phenotype of cross resistance between erythromycin and clindamycin. In Multilocus sequence typing (MLST), some commonly-detected resistance genes and virulence gene of these S. agalactiae isolates were investigated using polymerase chain reaction (PCR). Biofilms were detected by crystal violet staining. Our data demonstrated the correalation of the biofilm formation and OMA antimicrobial susceptibility of S.agalactiae clinical isolates with the carrier of virulence gene scpB. Conclusively, OMC exhibits the robust antimcirobial activity against clinical S. agalactiae isolates from China compared with DOX or MIN, and the carrier of the virulence gene scpB might correlate with the biofilm formation in OMC-resistant S. agalactiae.

Report on first confirmed outbreaks of contagious agalactia in small ruminants from Maharashtra State of India

Mycoplasma agalactiae causes contagious agalactia, a serious, economically important and OIE listed diseasesyndrome, in sheep and goats. There are no reports of this disease from Maharashtra state of India. The aim ofthe present investigation was to record epidemiological information and to detect M. agalactiae in clinical samplesfrom natural outbreaks of contagious agalactia. Clinically, anorexia, reluctance to follow the herd, lethargy and feverwere the initial clinical signs noted. Later, mastitis, arthritis and keratoconjunctivitis were noted as important clinicalsigns in both sheep and goats. However, all signs were not present in every case. DNA was extracted from mastiticmilk and articular exudate collected from suspected cases and PCR was performed using primers specific for 16S rRNA gene of Mycoplasma spp. The agarose gel electrophoresis of PCR product revealed the presence of bands of expected size (278 bp) indicating positive amplification. The nucleotide sequence analysis of PCR amplicons showed 100% identity with 16S rRNA gene of Mycoplasma agalactiae isolates JF4428 (Accession No. LT578418.1). The outbreaks of contagious agalactia were confirmed for the first time in Maharashtra state of India and carries epidemiological significance.

The prevalence, molecular characterization and antimicrobial resistance profiling of Streptococcus agalactiae isolated from clinical mastitis cases on large dairy farms in China.

This research communication aims to characterize the prevalence, molecular characterization and antimicrobial resistance profiling of Streptococcus agalactiae isolated from clinical mastitis in China. A total of 140 Strep. agalactiae isolates were identified from 12 out of 201 farms in 6 provinces, overall herd prevalence was 18.6% and the MLST analysis showed clonal complexes (CC) 103 and CC 67 were present in these herds with CC 103 predominant, accounting for 97.9%. Isolates were mostly sensitive to the tested antimicrobials: penicillin, ceftiofur, amoxi/clav, cefquinome, and vancomycin (100%), followed by cefalexin (97.9%), oxacillin (96.4%), enrofloxacin (95.7%), erythromycin (89.3%), and clindamycin (88.6%). Only 19.3 and 0.7% of isolates were sensitive to tetracycline and daptomycin, respectively, and sequence type (ST) 103 was most resistant to antimicrobials. In conclusion, CC 103 was the predominant subgroup of bovine mastitis Strep. agalactiae in China, and most antimicrobials apart from tetracycline and daptomycin were effective.

Natural outbreaks and molecular characteristics of Streptococcus agalactiae infection in farmed American bullfrog (Rana catesbeiana)

Between June and September 2019, a disease outbreak resulted in high mortality rates on four American bullfrog farms in Wenchang, Hainan Province, China. The main clinical signs included congestion and hemorrhage of the skin on the abdomen and hind limbs, ascites, hepatomegaly and internal organ hyperemia. Gram-positive cocci were isolated from the primary organs and tissues of 149 of 160 collected dead and moribund bullfrogs. The isolates were identified as Streptococcus agalactiae based on their morphology, biochemical characteristics and 16S rDNA sequencing analysis. S. agalactiae was isolated from diseased bullfrogs from all farms, and strain WH914 was selected for experimental infection of bullfrogs. Similar lesions were observed in the experimental bullfrogs, and 17.0%, 33.3% and 100.0% mortality occurred 14 days after administering 8.3 × 104 CFU/ml, 8.3 × 106 CFU/ml and 8.3 × 108 CFU/ml of the bacteria, respectively. Molecular serotyping of the capsular polysaccharide gene clusters revealed that the S. agalactiae isolates were serotype Ib. The sequence type (ST) of 30 isolates randomly selected from each bullfrog farm was determined by multilocus sequence typing as ST261. Virulence gene profiles for 14 virulence-related genes were constructed via multiplex PCR. All strains contained pavA, sip, iagA, cfb, hylB, bibA, cspA and fbsA but lacked cylE, bac, imb, scpB, rib and bca. S. agalactiae serotype Ib ST261 was isolated from sick or deceased farmed bullfrogs and is recognized as a pathogen of bullfrogs. Natural infections of S. agalactiae serotype Ib ST261 have not been previously reported in American bullfrogs.

Detection of virulence associated genes in Streptococcus agalactiae isolated from bovine mastitis.

Mastitis is one of the most expensive diseases in the dairy industry. It causes heavy monetary losses by decreasing milk production and treatment cost. Streptococcus agalactiae, the cause of contagious bovine mastitis, possesses various virulence factors that contribute to pathogenicity. The main aim of the study was to evaluate the distribution of virulence genes of S. agalactiae. In the current work, 98 Streptococcus species were isolated from 320 milk samples, collected from Veterinary Clinical Complex, Junagadh. Out of the isolates, 42 S. agalactiae isolates were used for virulence genes detection. All Streptococcus spp. were confirmed at genus level by targeting tuf gene, and S. agalactiae was identified at species level by targeting 16S rRNA gene. For virulence gene detection, scpB, cfb, and cylE genes were targeted. Of 42 S. agalactiae isolates, 15, 16, and 10 isolates possessed scpB, cfb, and cylE genes, respectively. This study aids us to know virulence characteristics and mechanisms responsible for the development of new strains in mastitis epidemiology in response to prevention and control strategies.

Prevalence, antimicrobial susceptibility, and genotyping of Streptococcus agalactiae in Tilapia fish (Oreochromis niloticus) in Egypt.

Objectives:Streptococcus agalactiae is a zoonotic human and animal pathogen that causes global economic losses in aquaculture and fatal outcomes in Tilapia. This study aimed to identify S. agalactiae isolated from different fish sources intended for human consumption phenotypically and genotypically and to characterize the virulence-associated genes fbsA (fibrinogen-binding protein FbsA), cfb (CAMP factor), and pbp1A/ponA (penicillin-binding protein 1A).Materials and Methods:Three hundred Nile Tilapia fish (Oreochromis niloticus) were collected from different farms and retail shops in Dakahlia and Damietta, Egypt, during the summer of 2020. The samples were examined using routine phenotypic methods, then characterized using polymerase chain reaction (PCR) targeting S. agalactiae-specific dltS gene. All S. agalactiae isolates were examined for the susceptibility to ten antimicrobial agents by the disc diffusion method. The virulence-associated genes (fbsA, cfb, and pbp1A/ponA) were characterized using multiplex-PCR.Results:Streptococcus agalactiae was detected in 7% (n = 21/300) samples. The isolates showed high resistance against ampicillin and erythromycin (20/21; 95%) for each. The most predominant antibiotypes through isolates were P, CN, SXT, CRO, TE, CTX, E, AMP, at 10.5% for each antibiotype. A total of 19 (90.5%) of S. agalactiae isolates showed multi-drug resistance (MDR), and those were recovered from market Tilapia fish. The virulence-associated genes (fbsA, cfb, and pbp1A/ponA) were identified in the S. agalactiae as 100%, 76%, and 52%, respectively.Conclusions:The MDR S. agalactiae detected in raw Tilapia fish pose a significant health hazard to consumers due to their zoonotic characteristics.

Multiplex PCR method for detection of variability in genes encoding the alpha proteins of Streptococcus agalactiae

The majority of Streptococcus agalactiae strains express one or more surface-anchored proteins that vary by strain. These proteins, which are characteristic for S. agalactiae, and are able to induce protective antibodies, include the alpha C, Rib proteins, alpha-like protein 2, and alpha-like protein 3. In this study was developed multiplex PCR method for detection of genes encoding these proteins, and its occurrence within a various S. agalactiae isolates of bovine origin. Also are reported two new genes from bovine isolates of S. agalactiae amplified by PCR, encoding other putative members of the family, alphalike protein 6, and alpha-like protein 7. They contain an overall genetic organization highly similar to that of the alpha C and Rib proteins.

Dog/cat-origin quinolone-resistant Streptococcus agalactiae isolates with point mutations in quinolone resistance-determining regions: Relatedness with clonal complex 10

ObjectiveWe aimed to investigate dog/cat-origin quinolone-resistant Streptococcus agalactiae isolates with point mutations in quinolone resistance-determining regions (QRDRs) and to define the relatedness between quinolone-resistant isolates and their microbiological features of capsular genotype, sequence type (ST)/clonal complex (CC), and antimicrobial resistance (AMR) gene. MethodsWith dog/cat-origin 22 isolates, type strain, and human-origin 6 isolates, we performed antimicrobial susceptibility testing by agar plate dilution method using levofloxacin, ciprofloxacin, and moxifloxacin. We also determined amino acid sequences in QRDRs of gyrA/gyrB/parC/parE genes and their point mutations. We conducted capsular genotyping, multilocus sequence typing, and AMR genotyping in our previous investigations. Correlations between quinolone-resistant population and their microbiological features were examined. ResultsWe found dog/cat-origin seven (31.8%) quinolone-resistant isolates harboring minimum inhibitory concentrations (MICs) of levofloxacin 16–32 μg/mL, ciprofloxacin 32 μg/mL, and moxifloxacin 2–4 μg/mL: human three isolates indicated MICs of levofloxacin 16–64 μg/mL, ciprofloxacin 32 μg/mL, and moxifloxacin 2–16 μg/mL. Point mutations Ser81Leu in gyrA and Ser79Phe/Ser79Tyr/Asp83Asn/Gly128Asp in parC were observed among these resistant isolates: mutations Leu495Ile/Val503Ile in parE was found among quinolone-nonresistant isolates. There was a significant correlation between dog/cat-origin quinolone-resistant population and ST10 (p = 0.023)/CC10 (p = 0.021). ConclusionTo our best knowledge, this is the first report assessing dog/cat-origin quinolone-resistant S. agalactiae. Our observations could be applied in future, by veterinarians while treating dogs and cats with clinical symptoms/signs suggestive of streptococcal infections.

Emergence of Multi-Drug Resistance and Its Association With Uncommon Serotypes of Streptococcus agalactiae Isolated From Non-neonatal Patients in Thailand.

Group B streptococcus (GBS) or Streptococcus agalactiae is an opportunistic pathogen that causes serious illness in newborns, pregnant women, and adults. However, insufficient detection methods and disease prevention programs have contributed to an increase in the incidence and fatality rates associated with this pathogen in non-neonatal patients. This study aimed to investigate factors of the observed increased incidence by investigation of serotype distribution, virulence factors, and antimicrobial susceptibility patterns from invasive GBS disease among non-neonatal patients in Thailand. During 2017–2018, a total of 109 S. agalactiae isolates were collected from non-pregnant patients. There were 62 males and 47 females, with an average age of 63.5 years (range: 20 – 96). Serotypes were determined by latex agglutination assay and multiplex polymerase chain reaction (PCR)-based assay. Among those isolates, seven virulence genes (rib, bca, pavA, lmb, scpB, cylE, and cfb) were detected by PCR amplification, and were determined for their susceptibility to 20 antimicrobial agents using a SensititreTM Streptococcus species STP6F AST plate. Among the study isolates, serotype III was predominant (52.3%), followed by serotype V and serotype VI (13.8% for each), serotype Ib (11.9%), and other serotypes (8.2%). Of the seven virulence genes, pavA was found in 67.0%. Except for one, there were no significant differences in virulence genes between serotype III and non-serotype III. Study isolates showed an overall rate of non-susceptibility to penicillin, the first-line antibiotic, of only 0.9%, whereas the resistance rates measured in tetracycline, clindamycin, azithromycin, and erythromycin were 41.3, 22.0, 22.0, and 22.0%, respectively. Strains that were resistant to all four of those drugs were significantly associated with non-serotype III (p < 0.001). Using multi-locus sequence typing (MLST), 40.0% of the four-drug-resistant isolates belonged to serotype VI/ST1, followed by serotype Ib/ST1 (35.0%). Cluster analysis with global GBS isolates suggested that the multiple drug-resistant isolates to be strongly associated with the clonal complex (CC) 1 (p < 0.001). Compared to the 2014 study of 210 invasive GBS isolates conducted in 12 tertiary hospitals in Thailand, the proportion of serotype III has dramatically dropped from nearly 90% to about 50%. This suggests that resistances to the second-line antibiotics for GBS might be the selective pressure causing the high prevalence of non-serotype III isolates.