Krecek R C, Maingi N
Department of Zoology and Entomology, Faculty of Natural and Agricultural Sciences, University of Pretoria, South Africa.
Vet Parasitol. 2004 Jul 14;122(3):233-44. doi: 10.1016/j.vetpar.2004.04.009.
A laboratory trial to determine the efficacy of two methods in recovering known numbers of third-stage (L3) strongylid nematode larvae from herbage was carried out. Herbage samples consisting almost entirely of star grass (Cynodon aethiopicus) that had no L3 nematode parasitic larvae were collected at Onderstepoort, South Africa. Two hundred grams samples were placed in fibreglass fly gauze bags and seeded with third-stage strongylid nematode larvae at 11 different levels of herbage infectivity ranging from 50 to 8000 L3/kg. Eight replicates were prepared for each of the 11 levels of herbage infectivity. Four of these were processed using a modified automatic Speed Queen heavy-duty washing machine at a regular normal cycle, followed by isolation of larvae through centrifugation-flotation in saturated sugar solution. Larvae in the other four samples were recovered after soaking the herbage in water overnight and the larvae isolated with the Baermann technique of the washing. There was a strong correlation between the number of larvae recovered using both methods and the number of larvae in the seeded samples, indicating that the two methods give a good indication of changes in the numbers of larvae on pasture if applied in epidemiological studies. The washing machine method recovered higher numbers of larvae than the soaking and Baermann method at all levels of pasture seeding, probably because the machine washed the samples more thoroughly and a sugar centrifugation-flotation step was used. Larval suspensions obtained using the washing machine method were therefore cleaner and thus easier to examine under the microscope. In contrast, the soaking and Baermann method may be more suitable in field-work, especially in places where resources and equipment are scarce, as it is less costly in equipment and less labour intensive. Neither method recovered all the larvae from the seeded samples. The recovery rates for the washing machine method ranged from 18 to 41% while those for the soaking and Baermann method ranged from 0 to 27%. Practical application of the two methods to estimate the number of nematode larvae on pastures without applying a correction factor would therefore result in a significant underestimation. This study provides a model, which can be applied in various laboratories to determine the larval recovery rates for techniques being used and the application of a correction factor when estimating the actual numbers of larvae on pasture.
开展了一项实验室试验，以确定两种方法从牧草中回收已知数量的第三期（L3）圆线虫幼虫的效果。在南非翁德斯特普特采集了几乎完全由无L3线虫寄生幼虫的非洲狗牙根（Cynodon aethiopicus）组成的牧草样本。将200克样本放入玻璃纤维蝇纱布袋中，并以每千克牧草50至8000个L3的11种不同感染水平接种第三期圆线虫幼虫。对11种牧草感染水平中的每一种都准备了8个重复样本。其中4个样本使用改良的自动Speed Queen重型洗衣机以常规正常程序处理，然后通过在饱和糖溶液中离心浮选分离幼虫。另外4个样本中的幼虫在将牧草浸泡过夜后回收，并使用洗涤的贝尔曼技术分离幼虫。两种方法回收的幼虫数量与接种样本中的幼虫数量之间存在很强的相关性，这表明如果应用于流行病学研究，这两种方法能很好地反映牧场上幼虫数量的变化。在所有牧草播种水平下，洗衣机法回收的幼虫数量均高于浸泡法和贝尔曼法，这可能是因为洗衣机对样本的清洗更彻底，且使用了糖离心浮选步骤。因此，使用洗衣机法获得的幼虫悬浮液更清洁，在显微镜下更容易检查。相比之下，浸泡法和贝尔曼法可能更适合野外工作，特别是在资源和设备匮乏的地方，因为它的设备成本较低且劳动强度较小。两种方法均未从接种样本中回收所有幼虫。洗衣机法的回收率为18%至41%，而浸泡法和贝尔曼法的回收率为0%至27%。因此，在不应用校正因子的情况下，将这两种方法实际应用于估计牧场上线虫幼虫的数量会导致显著低估。本研究提供了一个模型，可应用于各个实验室，以确定所使用技术的幼虫回收率，以及在估计牧场上幼虫实际数量时应用校正因子。
