Yersinia pestis: examining wildlife plague surveillance in China and the USA

Plague is a zoonotic disease caused by the bacterium Yersinia pestis Lehmann and Neumann, 1896. Although it is essentially a disease of rodents, plague can also be transmitted to people. Historically, plague has caused massive morbidity and mortality events in human populations, and has recently been classified as a reemerging disease in many parts of the world. This public health threat has led many countries to set up wild and domestic animal surveillance programs in an attempt to monitor plague activity that could potentially spill over into human populations. Both China and the USA have plague surveillance programs in place, but the disease dynamics differ in each country. We present data on plague seroprevalence in wildlife and review different approaches for plague surveillance in the 2 countries. The need to better comprehend plague dynamics, combined with the fact that there are still several thousand human plague cases per year, make well-designed wildlife surveillance programs a critical part of both understanding plague risks to humans and preventing disease outbreaks in the future.     

Adaptive strategies of African horse sickness virus to facilitate vector transmission

African horse sickness virus (AHSV) is an orbivirus that is usually transmitted between its equid hosts by adult Culicoides midges. In this article, we review the ways in which AHSV may have adapted to this mode of transmission. The AHSV particle can be modified by the pH or proteolytic enzymes of its immediate environment, altering its ability to infect different cell types. The degree of pathogenesis in the host and vector may also represent adaptations maximising the likelihood of successful vectorial transmission. However, speculation upon several adaptations for vectorial transmission is based upon research on related viruses such as bluetongue virus (BTV), and further direct studies of AHSV are required in order to improve our understanding of this important virus.     

Yersinia enterocolitica in sheep - a high frequency of biotype 1A

Background

Pigs are regarded as the main reservoir for human pathogenic Yersinia enterocolitica, which is dominated by bioserotype 4/O:3. Other animals, including sheep, have occasionally been reported as carriers of pathogenic strains of Y. enterocolitica. To our knowledge, this is the first study performed in the Nordic countries in which the presence of Y. enterocolitica in sheep is investigated.

Methods

Tonsils and faecal samples collected from sheep slaughtered on the island Gotland (Sweden) from September 2010 through January 2011 were analysed for presence of Y. enterocolitica. In an attempt to maximize recovery, several cultural strategies were applied. Various non-selective media were used and different temperatures and durations of the enrichment were applied before subculturing on Cefsulodin Irgasan Novobiocin (CIN) agar. Presumptive Y. enterocolitica colonies were subjected to urease, API 20E and agglutination test. Yersinia enterocolitica isolates were biotyped, serotyped, and tested for pathogenicity using a TaqMan PCR directed towards the ail-gene that is associated with human pathogenic strains of Y. enterocolitica.

Results

The samples collected from 99 sheep yielded 567 presumptive Y. enterocolitica colonies. Eighty urease positive isolates, from 35 sheep, were identified as Y. enterocolitica by API 20E. Thirty-four of 35 further subtyped Y. enterocolitica isolates, all from faecal samples, belonged to biotype 1A serotype O:5, O:6. O:13,7 and O:10. One strain was Yersinia mollaretii serotype O:62. No human pathogenic strains of Y. enterocolitica were found in the investigated sheep. Other species identified were Y. kristensenii (n = 4), Y. frederiksenii/intermedia (n = 3), Providencia rettgeri (n = 2), Serratia marcescens (n = 1) and Raoultella ornithinolytica (n = 1).

Conclusions

This study does not support the hypothesis that sheep play an important role in transmission of the known human pathogenic Y. enterocolitica in the studied geographical region. However, because there are studies indicating that some strains of Y. enterocolitica biotype 1A may cause disease in humans, the relative importance of sheep as carriers of human pathogenic strains of Y. enterocolitica remains unclear. Tonsils do not appear to be favourable sites for Y. enterocolitica biotype 1A in sheep.     

Local-scale diversity of Yersinia pestis: A case study from Ambohitromby,Ankazobe District,Madagascar

Plague is a re-emerging zoonotic disease and a major public health concern in several portions of the world, especially in Madagascar. We report on the presence of different subtypes of Yersinia pestis co-occurring in the same locality. After confirmation of a human plague case in Ambohitromby Commune (Ankazobe District) via isolation of Y. pestis, we undertook small mammal trapping to identify the circulation of Y. pestis amongst rodents in this locality; blood samples were collected from rodents for seroprevalence analysis. Of the 60 individuals of Rattus rattus captured, one yielded an isolate of Y. pestis, 13 others were positive for F1 antigen of Y. pestis using a rapid diagnostic test, and 4 were PCR positive targeting the caf1 and pla genes; 28/60 (46.7%) of the captured R. rattus were seropositive for Y. pestis. Whole-genome SNP analyses revealed that the two isolates obtained from the human case, and the R. rattus belonged to two different subtypes of Y. pestis (s05 and s13, respectively) that were circulating concurrently in Ambohitromby in 2016. Three Y. pestis subtypes (s03, s05 and s13) have now been isolated from Ambohitromby. Subtype s05 had been persisting there for >10 years but one or both of the other subtypes may have been introduced from the Central Highlands region as they were not observed in previous years (s13) or only observed once previously (s03). High seroprevalence against Y. pestis in R. rattus suggests that a portion of the local murine population may have acquired resistance to Y. pestis. Future research should focus on genomically characterizing Y. pestis strains circulating in Ankazobe District and other plague-endemic regions of Madagascar to better understand the overall phylogeography of Y. pestis.     

Yersiniosis caused by Yersinia pseudotuberculosis in captive toucans (Ramphastidae) and a Japanese squirrel (Sciurus lis) in zoological gardens in Japan

Two captive Keel-billed toucans and a Chestnut-mandibled toucan in another zoological garden died suddenlywithout any pre-existing symptoms, and three months later, a Japanese squirrel died of diarrhea. All theseanimals showed necrotic enteritis and multifocal necrosis in the liver and spleen with Gram negative bacilli.The bacilli showed strong positive immunolabeling for Yersinia pseudotuberculosis O4 in theKeel-billed toucans, Y. pseudotuberculosis O2 in the Chestnut-mandibled toucan and Y.pseudotuberculosis O1 in the Japanese squirrel, while Y. pseudotuberculosis 4b, 2band 1b were respectively isolated from the lesions. To our knowledge, this might be the first reported case offatal yersiniosis in a Japanese squirrel in the world as well as in toucans in Japan.     

Development and evaluation of a multiplex PCR assay for simultaneous detection of Flavobacterium psychrophilum, Yersinia ruckeri and Aeromonas salmonicida subsp. salmonicida in culture fisheries

Bacterial cold water disease, enteric red mouth disease and frunculosis are the common bacterial diseases of fish worldwide. The etiologic agents of these diseases are Flavobacterium (F.) psychrophilum, Yersinia (Y.) ruckeri and Aeromonas (A.) salmonicida subsp. salmonicida, respectively. In this study, a multiplex polymerase chain reaction (m-PCR) method with YER8/10-Fer3/4-FP1/3 primer pairs which can identify these fish pathogens simultaneously was developed and optimized. In optimized conditions, neither false specific nor nonspecific amplification occurred. The detection limits of the m-PCR method using DNA extracts from dilutions of pure cultures of bacteria were 35 pg for Y. ruckeri and F. psychrophilum and 70 pg for A. salmonicida subsp. salmonicida. It was determined that 15 CFU Y. ruckeri and F. psychrophilum and 30 CFU A. salmonicida subsp. salmonicida could be detected by m-PCR developed using genomic DNA extracted from dilutions of the suspensions. The detection limits in the presence of tissue debris were 125 CFU for Y. ruckeri and F. psychrophilum and 250 CFU for A. salmonicida subsp. salmonicida. In conclusion, we submit that the m-PCR method developed and optimized in this study can be used for accurate and rapid identification of these bacteria.     

鱼源小肠结肠炎耶尔森氏菌的耐药性研究

本试验采用聚合酶链反应(PCR)方法检测鱼源小肠结肠炎耶尔森氏菌的四环素类耐药基因(tetA、tetC、tetM)、磺胺类耐药基因(sul1、sul2、sul3)和氨基糖苷类耐药基因(aph(3')-Ⅲa、aac(6')-Ⅰb、ant(3')-Ⅰa、aac(3)-Ⅱa),并采用κ-B法检测该菌对14种抗生素的耐药表型。结果表明,小肠结肠炎耶尔森氏菌8种耐药基因(tetC、tetM、sul1、sul2,aph(3')-Ⅱa、aac(6')-Ⅰb、ant(3')-Ⅰa、aac(3)-Ⅱa)被检测出,而tetA、sul3基因未被检测出。该菌株对复方新诺明、磺胺异恶唑、红霉素、利福平、洁霉素和头孢噻吩6种抗生素耐药,对氟哌酸、氟苯尼考和头孢曲松等8种抗生素敏感。这为治疗小肠结肠炎耶尔森氏菌引起的鱼病积累了资料。     

Ecological fitness and strategies of adaptation of Bartonella species to their hosts and vectors

Bartonella spp. are facultative intracellular bacteria that cause characteristic host-restricted hemotropic infections in mammals and are typically transmitted by blood-sucking arthropods. In the mammalian reservoir, these bacteria initially infect a yet unrecognized primary niche, which seeds organisms into the blood stream leading to the establishment of a long-lasting intra-erythrocytic bacteremia as the hall-mark of infection. Bacterial type IV secretion systems, which are supra-molecular transporters ancestrally related to bacterial conjugation systems, represent crucial pathogenicity factors that have contributed to a radial expansion of the Bartonella lineage in nature by facilitating adaptation to unique mammalian hosts. On the molecular level, the type IV secretion system VirB/VirD4 is known to translocate a cocktail of different effector proteins into host cells, which subvert multiple cellular functions to the benefit of the infecting pathogen. Furthermore, bacterial adhesins mediate a critical, early step in the pathogenesis of the bartonellae by binding to extracellular matrix components of host cells, which leads to firm bacterial adhesion to the cell surface as a prerequisite for the efficient translocation of type IV secretion effector proteins. The best-studied adhesins in bartonellae are the orthologous trimeric autotransporter adhesins, BadA in Bartonella henselae and the Vomp family in Bartonella quintana. Genetic diversity and strain variability also appear to enhance the ability of bartonellae to invade not only specific reservoir hosts, but also accidental hosts, as shown for B. henselae. Bartonellae have been identified in many different blood-sucking arthropods, in which they are typically found to cause extracellular infections of the mid-gut epithelium. Adaptation to specific vectors and reservoirs seems to be a common strategy of bartonellae for transmission and host diversity. However, knowledge regarding arthropod specificity/restriction, the mode of transmission, and the bacterial factors involved in arthropod infection and transmission is still limited.     

云南省2株鹿流行性出血热病毒的分离鉴定

为了监测云南省反刍动物鹿流行性出血热病毒(epizootic hemorrhagic disease virus,EHDV)的感染情况,本试验在云南省师宗县设立了监控点,筛选出EHDV抗体阴性的10头牛和5只羊作为哨兵动物,每年的5～10月份每周采血1次,11月到次年4月份每月采血1次,通过酶联免疫吸附试验监测其抗体转阳情况,用转阳牛的红细胞接种BHK以分离病毒,用病毒RT-PCR和中和试验鉴定病毒。结果显示,从2头转阳牛的抗凝血中分离到2株EHDV,其TCID₅₀分别为10^-2.5/0.1 mL和10^-3.44/0.1 mL,血清型均为EHDV-5型。本试验在云南省分离到EHDV,明确了云南省反刍动物存在EHDV感染,为我国监控EHDV流行情况及疫病风险防范提供了重要借鉴意义。     

Acute toxoplasmosis in three wild arctic foxes (Alopex lagopus) from Svalbard; one with co-infections of Salmonella Enteritidis PT1 and Yersinia pseudotuberculosis serotype 2b

Acute disseminated toxoplasmosis was diagnosed in three wild arctic foxes (Alopex lagopus) that were found dead in the same locality on Svalbard (Norway). The animals included one adult female and two 4-months-old pups. The adult fox was severely jaundiced. Necropsy revealed multifocal, acute, necrotizing hepatitis, acute interstitial pneumonia, and scattered foci of brain gliosis, often associated with Toxoplasma tachyzoites. One pup also had Toxoplasma-associated meningitis. In addition, the latter animal was infected with Yersinia pseudotuberculosis serotype 2b and Salmonella Enteritidis phage type 1 (PT1), which may have contributed to the severity of the Toxoplasma infection in this animal. The diagnosis of toxoplasmosis was confirmed by positive immunohistochemistry and detection of anti-Toxoplasma gondii antibodies in serum of all foxes. The animals were negative for Neospora caninum, canine distemper virus, canine adenovirus, and rabies virus on immunolabelling of tissue sections and smears.     

Spatial analysis of Yersinia pestis and Bartonella vinsonii subsp. berkhoffii seroprevalence in California coyotes (Canis latrans)

Zoonotic transmission of sylvatic plague caused by Yersinia pestis occurs in California, USA. Human infections with various Bartonella species have been reported recently. Coyotes (Canis latrans) are ubiquitous throughout California and can become infected with both bacterial agents, making the species useful for surveillance purposes. This study examined the geographic distribution of 863 coyotes tested for Y. pestis and Bartonella vinsonii subsp. berkhoffii serologic status to gain insight into the natural history of B. vinsonii subsp. berkhoffii and to characterize the spatial distribution of the two agents.

We found 11.7% of specimens positive to Y. pestis and 35.5% positive to B. vinsonii subsp. berkhoffii. The two pathogens had distinct spatial clusters: Y. pestis was more prevalent in eastern portions of the state and B. vinsonii subsp. berkhoffii in coastal regions. Prevalence of Y. pestis increased with increasing elevation, whereas prevalence of B. vinsonii subsp. berkhoffii decreased with increasing elevation. There were differences in the proportions of positive animals on a yearly basis to both pathogens.     

Changes in various metabolic parameters in blood and milk during experimental Escherichia coli mastitis for primiparous Holstein dairy cows during early lactation

Background

The objective of this study was to characterize the changes in various metabolic parameters in blood and milk during IMI challenge with Escherichia coli (E. coli) for dairy cows during early lactation. Thirty, healthy primiparous Holstein cows were infused (h = 0) with ~20-40 cfu of live E. coli into one front mammary quarter at ~4-6 wk in lactation. Daily feed intake and milk yield were recorded. At –12, 0, 3, 6, 12, 18, 24, 36, 48, 60, 72, 96, 108, 120, 132, 144, 156, 168, 180 and 192 h relative to challenge rectal temperatures were recorded and quarter foremilk was collected for analysis of shedding of E. coli. Composite milk samples were collected at -180, -132, -84, -36, -12, 12, 24, 36, 48, 60, 72, 84, 96, 132 and 180 h relative to challenge (h = 0) and analyzed for lactate dehydrogenase (LDH), somatic cell count, fat, protein, lactose, citrate, beta-hydroxybutyrate (BHBA), free glucose (fglu), and glucose-6-phosphate (G6P). Blood was collected at -12, 0, 3, 6, 12, 18, 24, 36, 60, 72, 84, 132 and 180 h relative to challenge and analyzed for plasma non-esterified fatty acids (NEFA), BHBA and glucose concentration. A generalized linear mixed model was used to determine the effect of IMI challenge on metabolic responses of cows during early lactation.

Results

By 12 h, E. coli was recovered from challenged quarters and shedding continued through 72 h. Rectal temperature peaked by 12 h post-challenge and returned to pre-challenge values by 36 h post-IMI challenge. Daily feed intake and milk yield decreased (P <0.05) by 1 and 2 d, respectively, after mastitis challenge. Plasma BHBA decreased (12 h; P <0.05) from 0.96 ± 1.1 at 0 h to 0.57 ± 0.64 mmol/L by 18 h whereas concentration of plasma NEFA (18 h) and glucose (24 h) were significantly greater, 11 and 27%, respectively, after challenge. In milk, fglu, lactose, citrate, fat and protein yield were lower whereas yield of BHBA and G6P were higher after challenge when compared to pre-challenge values.

Conclusions

Changes in metabolites in blood and milk were most likely associated with drops in feed intake and milk yield. However, the early rise in plasma NEFA may also signify enhanced adipose tissue lipolysis. Lower concentrations of plasma BHBA may be attributed to an increase transfer into milk after IMI. Decreases in both milk lactose yield and % after challenge may be partly attributed to reduced conversion of fglu to lactose. Rises in G6P yield and concentration in milk after challenge (24 h) may signify increased conversion of fglu to G6P. Results identify changes in various metabolic parameters in blood and milk after IMI challenge with E. coli in dairy cows that may partly explain the partitioning of nutrients and changes in milk components after IMI for cows during early lactation.     

In vitro susceptibilities of Leptospira spp. and Borrelia burgdorferi isolates to amoxicillin, tilmicosin, and enrofloxacin

Antimicrobial susceptibility testing was conducted with 6 different spirochetal strains (4 strains of Leptospira spp. and 2 strains of Borrelia burgdorferi) against 3 antimicrobial agents, commonly used in equine and bovine practice. The ranges of MIC and MBC of amoxicillin against Leptospira spp. were 0.05-6.25 µg/ml and 6.25-25.0 µg/ml, respectively. And the ranges of minimal inhibitory concentration (MIC) and minimal bactericidal concentration (MBC) of amoxicillin against B. burgdorferi were 0.05-0.39 µg/ml and 0.20-0.78 µg/ml, respectively. The ranges of MIC and MBC of enrofloxacin against Leptospira spp. were 0.05-0.39 µg/ml and 0.05-0.39 µg/ml, respectively. Two strains of B. burgdorferi were resistant to enrofloxacin at the highest concentration tested for MBC (≥100 µg/ml). Therefore, the potential role of tilmicosin in the treatment of leptospirosis and borreliosis should be further evaluated in animal models to understand whether the in vivo studies will confirm in vitro results. All spirochetal isolates were inhibited (MIC) and were killed (MBC) by tilmicosin at concentrations below the limit of testing (≤0.01 µg/ml).     

An evaluation of Irish cattle herds with inconclusive serological evidence of bovine brucellosis

Since 1998, there has been a steady decline in herd restrictions and de-populations in Ireland due to bovine brucellosis. There is concern that the interpretation of laboratory results may become increasingly problematic, as brucellosis prevalence falls in Ireland. Therefore, the purpose of the current study was to evaluate the infection status of Irish herds and animals with inconclusive serological evidence of bovine brucellosis. During 12 months from September 1, 2004, laboratory and observational epidemiological data were collected from all Irish herds where animal testing identified at least one animal with a complement fixation test (CFT) reading greater than zero and/or a positive result to the indirect enzyme-linked immunosorbent assay (iELISA). Due to the observational nature of the study, we have robust estimates of the relative, but not the absolute, performance of the CFT, iELISA and brucellin skin test (BST). Herds were divided into three categories (Group A, B or C) on the basis of test results at initial assessment. A total of 639 herds were enrolled into the study, and observed for at least two years following enrolment. A rising CFT titre, with a CFT reading of 111 International CFT Units (IU) or greater at the subsequent blood test, was generally associated with herds where other evidence of infection was also available. Knowledge of the CFT reading at the initial and a subsequent blood test proved useful in distinguishing false-positive and true-positive brucellosis results. There was poor correlation between the CFT and iELISA results, and between the CFT and BST results. As a result of this study, national policy has been modified to include re-sampling of all animals with CFT readings of 20 IU or greater. This project has also led to a reduction in the number of herds restricted, as well as restriction duration. It has also contributed to a reduction in the number of herds listed for contiguous tests, and therefore the potential for contiguity testing of false positive results.     

Prevalence of bacterial genotypes and outcome of bovine clinical mastitis due to Streptococcus dysgalactiae and Streptococcus uberis

Background

Streptococcus dysgalactiae and Streptococcus uberis are common causes of clinical mastitis (CM) in dairy cows. In the present study genotype variation of S. dysgalactiae and S. uberis was investigated, as well as the influence of bacterial species, or genotype within species, on the outcome of veterinary-treated CM (VTCM). Isolates of S. dysgalactiae (n = 132) and S. uberis (n = 97) were genotyped using pulsed-field gel electrophoresis. Identical banding patterns were called pulsotypes. Outcome measurements used were cow composite SCC, milk yield, additional registered VTCMs and culling rate during a four-month follow-up period.

Results

In total, 71 S. dysgalactiae pulsotypes were identified. Nineteen of the pulsotypes were isolated from more than one herd; the remaining pulsotypes were only found once each in the material. All S. uberis isolates were of different pulsotypes. During the follow-up period, the SCC of S. dysgalactiae-cows was significantly lower than the SCC of S. uberis-cows (P <0.05). Median SCC of S. dysgalactiae-cows was 71 500 cells/ml and of S. uberis-cows 108 000 cells/ml. No other differences in outcome parameters could be identified between species or genotypes.

Conclusions

Identical S. dysgalactiae genotypes were isolated from more than one herd, suggesting some spread of this pathogen between Swedish dairy herds. The genetic variation among S. uberis isolates was substantial, and we found no evidence of spread of this pathogen between herds. The milk SCC was lower during the follow-up period if S. dysgalactiae rather than S. uberis was isolated from the case, indicating differences in treatment response between bacterial species.

Electronic supplementary material

The online version of this article (doi:10.1186/s13028-014-0080-0) contains supplementary material, which is available to authorized users.     

Initial adherence of EPEC,EHEC and VTEC to host cells

Initial adherence to host cells is the first step of the infection of enteropathogenic Escherichia coli (EPEC), enterohaemorrhagic Escherichia coli (EHEC) and verotoxigenic Escherichia coli (VTEC) strains. The importance of this step in the infection resides in the fact that (1) adherence is the first contact between bacteria and intestinal cells without which the other steps cannot occur and (2) adherence is the basis of host specificity for a lot of pathogens. This review describes the initial adhesins of the EPEC, EHEC and VTEC strains. During the last few years, several new adhesins and putative colonisation factors have been described, especially in EHEC strains. Only a few adhesins (BfpA, AF/R1, AF/R2, Ral, F18 adhesins) appear to be host and pathotype specific. The others are found in more than one species and/or pathotype (EPEC, EHEC, VTEC). Initial adherence of EPEC, EHEC and VTEC strains to host cells is probably mediated by multiple mechanisms.