Prototheca zopfii genotypes isolated from cow barns and bovine mastitis in Japan

This study is the first investigation on Japanese isolates of Prototheca zopfii from bovine mastitis and the cow-barn surroundings by molecular characterization to clarify routes of infection for bovine protothecal mastitis. We performed isolation of Prototheca from cow-barn surroundings (drinking water, sewage and feces) and milk samples from cases of bovine mastitis. Genotypes of the 32 isolates of P. zopfii from cow-barn surroundings and 67 isolates from mastitis were analyzed by genotype-specific PCR assays and restriction fragment length polymorphism (RFLP) assays. All mastitis isolates were identified as P. zopfii genotype 2. Conversely, 29 isolates from cow-barn surroundings were identified as P. zopfii genotypes 1 and 3 isolates as genotype 2, respectively. Given these results, both genotypes of P. zopfii could exist in cow-barn surroundings, but no sites were identified as frequent sources of P. zopfii genotype 2. P. zopfii isolates should thus be further explored with regard to genotype to clarify the reservoir of etiological agents in bovine Prototheca mastitis.     

Yeasts and algae isolated from cows with mastitis in the south-eastern part of Poland

The aim of the study was to evaluate the occurrence of mycotic and protohecal mastitis in herds in south-eastern part of Poland. A total of 3091 milk samples from udder quarters with clinical and subclinical mastitis from 29 dairy herds was investigated in this survey. Milk samples were plated as soon as possible on blood agar (BA), Mac Conkey agar, aesculin-tallium acetate-crystal violet agar, and Sabouraud agar. A hundred and thirty one yeast (4.23%) and eleven Protoheca zopfii (0.35%) strains were isolated from cows with clinical and subclinical mastitis. All the isolated fungi were the yeast classified into 4 genera (Candida, Trichosporon, Saccharomyces and Rhodotorula). The most frequently isolated yeasts were Candida sp., C. kefyr, C. humicola, C. rugosa and C. inconspicua. Both fungi and algae were isolated first of all during a confinement-housing season.     

Eradication of Prototheca zopfii infection in a dairy cattle herd

Protothecosis is a severe form of mastitis in dairy cows caused by colorless algae of the genus Prototheca. Since P. zopfii is highly resistant to all known chemotherapeutics, infected cows must be removed from the herd. Eradication measures are difficult since many chronically infected cows may become intermittent shedders. Therefore, cultural methods are insufficient for control measures. In order to eradicate Prototheca zopfii-mastitis in dairy cattle herds, two isotype specific indirect ELISA for detection of IgA and IgG1 in whey were used in a dairy herd highly affected with protothecal mastitis. All cows (n = 313) were tested four times in intervals of six months. Milk specimens were examined in parallel by cultivation and serologically using two indirect ELISA systems for specific IgA and IgG1 in whey. Cows tested Prototheca positive were consequently separated from the herd and slaughtered. At the first examination, 15.6% of the animals were found positive by culture, and 23.3% were positive in at least one of the ELISA systems. Within two years, protothecal prevalence and incidence decreased to zero indicating that the eradication strategy used was successful. In summary, serological identification of P. zopfii-infected lactating cows is an useful tool to eradicate protothecal bovine mastitis in infected herds.     

Prototheca zopfii mastitis in a herd of dairy cows

Mastitis caused by the colourless alga Prototheca zopfii was diagnosed in 17 of 120 cows in a dairy herd. Infection occurred in animals varying from 3-14 years old and was present in one to four quarters of each cow. Nine cases were associated with clinical mastitis characterised by the presence in milk of flakes or small clots. Somatic cell counts consistent with subclinical mastitis (>500 x 10(3) cells/ml) were recorded in five of the eight remaining cows. Histological examination of udder tissue showed the presence of granulomatous lesions associated with the presence of Prototheca. The problem was identified and controlled by repeated microbiological examination of milk samples from all lactating cows and immediate culling of infected animals. P. zopfii was also recovered from environmental water samples on this farm. It is suggested that infection may have occurred as a result of teat sores caused by trauma from a milking machine, and the tendency for cows to lay down on a race, the surface of which was sometimes flooded by drain water in which Prototheca were present.     

Bovine mastitis caused by Prototheca zopfii: a case study

An outbreak of severe indurative mastitis in newly calved cows, from which Prototheca zopfii was isolated, is described. The refractory nature of this type of mastitis and the ineffectiveness of the presently available intramammary preparations are highlighted.     

奶牛中型无绿藻性乳房炎

目前已有12个国家报道无绿藻性奶牛乳房炎,造成严重的经济损失并对公共安全存在潜在的危害.2010年春季,北京地区某奶牛场暴发中型无绿藻引起的临床型乳房炎.从患有临床型乳房炎的10头奶牛的11个乳区分离到8株酵母样微生物,鉴定为中型无绿藻.用16种抗生素药敏纸片进行药敏试验,结果显示中型无绿藻仅对庆大霉素、链霉素和阿米卡星等氨基糖苷类抗生素敏感.同化试验证明该分离株能发酵葡萄糖和丙三醇.本试验在我国首次报道了中型无绿藻能引起奶牛临床型乳房炎.     

Incidence and types of clinical mastitis in dairy herds with high and low somatic cell counts

Eighteen dairy herds were studied, 12 with a 12-month Dairy Herd Improvement Association herd mean somatic cell count (SCC) less than or equal to 150,000 cells/ml (low SCC) and 6 with a 12-month mean SCC greater than 700,000 cells/ml (high SCC). At the outset of the study, quarter samples for bacteriologic culture were collected (in duplicate) from all quarters of all lactating cows (whole herd culture). Subsequently, quarter milk samples for culture from all cows with clinical mastitis were collected for a period of 6 months. In the herds with low SCC, results of whole herd culture revealed low prevalence of intramammary infection attributable to all major pathogens (less than 4% of all quarters). Prevalence of infection with Streptococcus agalactiae (22.2% of all quarters) and Staphylococcus aureus (6.6% of all quarters) was significantly (P less than 0.05) higher in the herds with high SCC. Mean incidence of clinical mastitis in the herds with low SCC was 4.23 infections/100 cows/month (range, 0.42 to 10.25 infections). In the herds with high SCC, mean incidence was 2.91 infections/100 cows/month (range, 1.33 to 3.92 infections). In the herds with low SCC, infection type, as mean percentage of total clinically infected quarters sampled for culture/herd, was 0.0%, 2.2%, 12.3%, 43.5%, and 28.6% for Str agalactiae, S aureus, streptococci other than Str agalactiae, coliforms, and organisms not isolated, respectively. Respective percentages for the herds with high SCC were 41.5%, 18.3%, 12.6%, 8.0%, and 8.8%. During the study period (from April through January), incidence of clinical mastitis and clinical mastitis caused by coliform bacteria were highest in July and August for herds with low SCC.(ABSTRACT TRUNCATED AT 250 WORDS)     

Natural and experimental bovine intramammary infection with Prototheca zopfii

A naturally occurring case of chronic mastitis caused by Prototheca zopfii was detected in a dairy cow. The cow was observed for 12 weeks, and milk samples were cultured each week. Milk production was decreased, and high numbers (greater than 10(3)/ml) of the organism were detected in the milk. Six glands of 2 cows were inoculated with small numbers of P zopfii. All 3 cows were euthanatized, and at postmortem, a progressive pyogranulomatous inflammation was observed in all infected mammary glands. The organism was recovered from tissue and secretions from all inoculated glands and from lymph nodes draining these glands but not from other tissues.     

Review of the microbiological, pathological, and clinical aspects of bovine mastitis caused by the alga Prototheca zopfii

The mastitis caused by P. zopfii alga is a disease of high-producing, machine-milked dairy cows. It occurs worldwide in tropical and temperate climatic areas, and mostly appears sporadically in a therapy-resistant form. However, in poorly managed dairy herds it may be endemic, causing serious economic losses as a result of decreased milk quality and quantity and culling of infected animals. The biological properties of this pathogenic alga, the laboratory methods available for its isolation and identification, the pathological and clinical features of this form of mastitis, and the principles of its control are reviewed in this paper.     

Isolation of Mycoplasma bovis from bovine clinical mastitis cases in Northern Greece

Mycoplasma bovis was detected in 18/219 (8.2%) quarter milk samples collected from cases of bovine clinical mastitis in Northern Greece between November 1997 and March 1999. The cases occurred in 2/37 (5.4%) of the herds examined. The micro-organism was isolated from bulk milk samples (BTS) from the two positive herds but was not isolated from 111 composite milk samples collected from clinically healthy cows from all 37 herds. Isolates were identified as M. bovis by polymerase chain reaction (PCR) assay. Other micro-organisms were also isolated from the M. bovis positive samples. The M. bovis-positive cows had all been imported into Greece from other European countries.     

Comparison of the efficacy of cloxacillin alone and cloxacillin combined with an internal teat sealant for dry-cow therapy

All the quarters in the cows with high somatic cell counts in 10 herds were treated at drying off with either 600 mg cloxacillin or 600 mg cloxacillin and 4 g of an internal teat sealant containing 65 per cent bismuth subnitrate. The quarters were sampled daily for bacteriological tests for the three days before drying off and twice after calving to establish whether they were infected. Clinical mastitis cases were monitored from drying off until 100 days after calving. The odds of a quarter being bacteriologically negative after calving or developing clinical mastitis in the first 100 days after calving were investigated by multilevel logistic regression. The quarters treated with the internal sealant and cloxacillin were significantly more likely to be bacteriologically negative in the immediate period after calving and were significantly less likely to suffer clinical mastitis during the first 100 days after calving than the quarters treated with cloxacillin alone. There was more variation between cows than between herds in the underlying risk of an infection after calving, but there was more variation between herds than between cows in the underlying risk of clinical mastitis during the 100 days after calving.     

A field study of Mycoplasma bovis infection in cattle

After an outbreak of mastitis in cattle caused by Mycoplasma bovis a study was made in 5 herds with recent cases (principal herds) and in 4 control herds. In the principal herds, M. bovis was isolated from milk samples, nasal swabs, and from one vaginal swab. M. bovis was also isolated from nasal swabs of calves in 2 of the 4 control herds, whereas all milk samples and vaginal swabs from the control herds were negative. Evaluation of serum antibody titres to M. bovis among non-mastitic animals of 3 principal herds and 1 control herd showed no difference in distribution of the titre values, which generally were low. However, cows excreting M. bovis in the milk had high antibody titres. The way of introduction to the herds and the spread of the infection within the herds could not be established by the study, which was supplemented by a DNA restriction fragment analysis of a number of M. bovis isolates.     

Within herd comparison of teat dipping and dry cow therapy with only selective dry cow therapy in six herds

A within herd comparison of teat dipping and dry cow therapy (full treatment) with only selective dry cow therapy (partial treatment) was carried out in six commercial dairy herds for a two year period. In four herds, the incidence of clinical mastitis was 2 to 12 per cent higher in the partial treatment group. In another herd, in which the pattern of clinical mastitis isolates was unusual in that minor pathogens were isolated from 30 per cent of mastitis cases, the incidence was 43 per cent higher in the partial treatment group. In the remaining herd the incidence was 10 percent higher in the full treatment group. Streptococcus uberis mastitis was more common in the partial treatment groups of five herds; coliform mastitis was more common in the full treatment groups of two herds and similar in both groups in the other herds. High rates of coliform mastitis were associated with poor herd environmental conditions but this was not true for Strep uberis mastitis. Rates of staphylococcus aureus and Strep dysgalactiae mastitis were low in all herds. The level of major pathogen infection in cows completing the trial in all herds increased in the partial treatment group from 5 per cent of quarters at the start to 12 per cent at the finish of the trial. In the full treatment group, however, there was only a small increase in this level. In contrast, levels of Corynebacterium bovis infection increased by 17 per cent in both treatment groups. Continued use of teat dipping and dry cow therapy was associated with a higher rate of coliform mastitis in two of the three herds where there were poor standards of hygiene and husbandry.     

Aetiology of clinical mastitis in six Somerset dairy herds.

Clinical mastitis was monitored in six Somerset dairy herds for one year. The herds all had three-month geometric mean bulk milk somatic cell counts of less than 250,000 cells/ml. Escherichia coli was the predominant pathogen isolated on all the farms and in all months of the year. Environmental pathogens accounted for 61.4 per cent of all cases of clinical mastitis and for 79.3 per cent of the mastitis cases in which an aetiological agent was identified. The mean annual incidence was 41.6 cases per 100 cows (range 14 to 75). Affected cows suffered a mean of 1.5 cases and 16.4 per cent of quarters suffered at least one repeat case. Mastitis due to E. coli was more severe than mastitis due to other causes and it tended to be more severe in early lactation and during the housing period. Mastitis was significantly more severe (grades 2 and 3) in the herd with the lowest bulk milk somatic cell count and in the herd which was kept indoors throughout the year than in the other four herds. Mastitis was fatal in 2.2 per cent of cases and resulted in the death of 0.6 per cent of the lactating cows.     

Genotyping of bovine Prototheca mastitis isolates from Poland

Bovine mastitis due to unicellular, achlorophyllous algae of the genus Prototheca is a serious and complex ailment that accounts for high economic losses in the dairy industry. Bovine protothecal mastitis has been almost exclusively associated with only one species Prototheca zopfii. Recently, based on the 18S rDNA sequence analysis, P. zopfii has been divided into three genotypes (1-3), of which P. zopfii genotype 3 has been given the status of a new species Prototheca blaschkeae sp. nov. The aim of this study was to investigate the genotypic composition of the population of P. zopfii bovine mastitis isolates from Poland, according to that threefold classification. Of the 44 isolates tested, 43 (98%) were identified as P. zopfii genotype 2. Only one isolate was assigned to the newly established P. blaschkeae species. These findings clearly show the predominance of the P. zopfii genotype 2 in the etiology of bovine mammary protothecosis in Poland, but also underline the involvement of P. blaschkeae in the disease. The study is the first to provide molecular characterization of Polish P. zopfii mastitis isolates, collected from different regions of Poland.     

Clinical and bacteriological response to treatment of clinical mastitis with one of three intramammary antibiotics

AIM: To compare the proportions of clinical and bacteriological cure of glands of dairy cows diagnosed with clinical mastitis, following treatment with one of three different intramammary antibiotic preparations. METHODS: Cows from dairy cow herds (n=28) across New Zealand which were diagnosed with clinical mastitis in one or more glands at any stage of lactation were randomly assigned at the cow level within sequentially presented groups of three animals to be treated with one of three intramammary antibiotics. The treatments were 1 g procaine penicillin, 0.25 g cefuroxime, and a combination of 1 g procaine penicillin and 0.5 g dihydrostreptomycin (DHS). All treatments were infused on three occasions at 12-hourly intervals. Duplicate milk samples were collected for bacteriology before initial treatment, and 21-42 days later. Logistic regression or generalised linear mixed models were used to analyse the proportion of cows or quarters retreated for mastitis within 30 days of initial treatment ('clinical treatment failure'), and the proportion of glands from which bacteria were isolated initially but from which the same bacterial species was not re-isolated ('bacteriological cure'). RESULTS: The annual herd average cumulative incidence rate of clinical mastitis was 12.7 cases/100 cows. The incidence rate was higher in young (2-year-old) and old (> or = 9-year-old) cows relative to 3- and 4-year-old cows, and was higher in Friesian than Jersey or crossbred cows. Streptococcus uberis was the pathogen most commonly isolated, and its relative prevalence declined with time postpartum. Cows treated with cefuroxime were more likely (p<0.01) to be re-treated for clinical mastitis in the 30 days after initial treatment than cows treated with procaine penicillin or procaine penicillin and DHS. Bacteriological cure occurred in 74% of treated glands and there was no difference in the proportion of cures among the treatments (p=0.4). The proportion of cures was lower when treatment occurred 28-72 days after calving (p<0.01) and if a major pathogen was isolated (p<0.001). CONCLUSIONS AND CLINICAL RELEVANCE: There was no benefit in terms of clinical or bacteriological cure rate in treating clinical mastitis cases with the combination of procaine penicillin and DHS compared to treatment with procaine penicillin alone. The proportion of clinical mastitis cases re-treated differed among herds, and more cows treated with cefuroxime were retreated within 30 days of initial treatment. However, the bacteriological cure proportion was the same among the treatments. Cure proportions were lower in cows from which major mastitis pathogens were isolated and when treatment commenced 28-72 days after calving.     

Prevalence of clinical mastitis in 38 Waikato dairy herds in early lactation

AIM: To determine the prevalence of clinical mastitis in spring-calving dairy herds in the Waikato Region of New Zealand and to identify factors associated with variation in the prevalence of clinical mastitis between herds. METHOD: A total of 799 quarters from 595 dairy cows from 38 dairy herds were diagnosed by herd owners as having clinical mastitis between 8 July and 21 August 1997. Quarters diagnosed with clinical mastitis were sampled for bacterial culture and somatic cell count, and the presence of clots in the milk and the presence of udder oedema were assessed by a technician or veterinarian. RESULTS: Clinical mastitis was diagnosed in an average (+/-s.e.m.) of 9.9% (+/-0.8%, range 0.9-21.4%) of calved cows within the herds. Bacteria were not cultured from an average of 12.4 % (+/- 2.0%, range 0.0-45.5%) of cows and 22.3% (+/- 2.4%, range 0.0-54.0%) of quarters diagnosed as having clinical mastitis. There were significant differences between herds in the proportion of cows diagnosed with mastitis and in the proportion of clinical mastitis cases from which bacteria were not cultured. A decreased prevalence of clinical mastitis (p<0.001) was associated with an increased percentage of the herd treated with dry cow antibiotics. An increased prevalence of clinical mastitis (p<0.0001) was associated with both an increased percentage of cows treated in the previous season with lactating cow antibiotics and an increased percentage of heifers in the herd. Herds that were fed supplements before or during lactation had a higher prevalence of clinical mastitis than herds that were not fed supplements (p<0.001). An increased proportion of quarters diagnosed with clinical mastitis that did not culture bacteria was associated with an increased prevalence of clinical mastitis (p<0.001). The proportion of quarters that the technician or veterinarian found with evidence of clinical mastitis (i.e. a somatic cell count >500,000 cells/ml and the presence of either clots or udder oedema) within a herd was inversely related to the proportion of quarters within a herd from which no bacteria were isolated. CONCLUSION: There was a large variation in the prevalence of clinical mastitis and in the proportion of clinical quarters from which no bacteria were grown between herds. Management factors such as the use of dry cow therapy, feeding regimes and heifer replacement rates all affected the prevalence of clinical mastitis. Herd owners appear to differ in the sensitivity and specificity of their diagnosis of clinical mastitis, with bacteria not isolated from up to 50% of quarters diagnosed with clinical mastitis in some herds. Improvements in the specificity of herd owner diagnosis of clinical mastitis may reduce the use of antibiotics for mastitis during lactation and hence may reduce the risk of antibiotic contamination of milk supplied for human consumption.     

Mastitis prevalence and the selection of dry cow therapy programmes in dairy herds

Sales of intramammary cerates to a group of 142 dairy farms over a period of two years were monitored. On the assumption that three tubes of cerate are used to treat one case of mastitis, an estimate was made of the prevalence of clinical mastitis in these herds. The prevalence in seasonal supply herds was 21 cases per 100 cows per year and the prevalence in town supply herds was 66 cases per 100 cows per year. The usage of dry cow therapy was related to changes in the prevalence of clinical mastitis in these herds. The use of whole-herd therapy reduced the prevalence of clinical mastitis in herds which had a high prevalence in the first year, but in herds with a low prevalence of clinical mastitis the prevalence increased after the use of dry cow therapy.     

Post-milking teat dip use in dairy herds with high or low somatic cell counts.

Milk samples for bacteriologic culture were submitted from 71 dairy herds, 24 with low somatic cell count (SCC) and 47 with high SCC and high prevalence of subclinical mastitis. At the time of sample submission to the Mastitis Diagnostic Laboratory of Pennsylvania State University, information regarding the herd mastitis control practices was collected. A combined program of post-milking teat dipping (PMTD) and antibiotic treatment of all cows at the start of the nonlactating period was practiced more frequently for herds with low SCC, (P less than 0.001) than for herds with high SCC. Among all herds for which PMTD was practiced, a higher proportion (P less than 0.001) of those for which chlorhexidine-based products were used had low SCC than high SCC. Conversely, a higher proportion of herds for which a dip with an acrylic latex barrier was used had high SCC rather than low SCC (P = 0.002). For herds with high prevalence of subclinical mastitis, and despite a program of PMTD and treatment of all cows at the start of the nonlactating period, a change to a different germicidal teat dip product may be indicated to help reduce prevalence of infection.     

Characterization of Staphylococcus simulans strains isolated from cases of bovine mastitis

This study was conducted to characterize Staphylococcus simulans isolated from cases of bovine mastitis. A total of 134 isolates of S. simulans selected from 80 quarters from 61 cows or heifers in 37 different herds were characterized by EcoRI ribotyping. From 22 quarters two to seven consecutive isolates taken at weekly intervals were selected. Furthermore, three isolates from clinical infections in humans and two reference strains were included. A total of 16 different ribotypes were found, however, two types predominated. In most herds more than one type was found. From the 22 different quarters, where 76 paired or multiple isolates were at disposal, the same ribotype was constantly found in the same quarter. This study showed that S. simulans causing bovine mastitis could be divided into relatively large number of different types, but that two types predominated. More than one type could be found in the same herd and within different quarters of the same cow, but ribotyping confirmed that S. simulans could be the cause of persistent and stable infections.